Computational methods and theory for ion channel research.

Ion channels are fundamental biological devices that act as gates in order to ensure selective ion transport across cellular membranes; their operation constitutes the molecular mechanism through which basic biological functions, such as nerve signal transmission and muscle contraction, are carried out. Here, we review recent results in the field of computational research on ion channels, covering theoretical advances, state-of-the-art simulation approaches, and frontline modeling techniques. We also report on few selected applications of continuum and atomistic methods to characterize the mechanisms of permeation, selectivity, and gating in biological and model channels.

Biophysical modeling of ß-cells networks: Realistic architectures and heterogeneity effects.

The ß-cells dynamics is the regulator of insulin secretion in the pancreas, and its investigation is a central aspect in designing effective treatment strategies for diabetes. Despite great efforts, much is still unknown about the complex organization of such endocrine cells and realistic mathematical modeling represents a useful tool to elucidate key aspects of glucose control in humans. In this contribution, we study the human ß-cells collective behaviour, by modeling their electric and metabolic coupling in a cluster, of size and architecture similar to human islets of Langerhans. We focus on the effect of coupling on various dynamics regimes observed in the islets, that are spiking and bursting on multiple timescales. In particular, we test the effect of hubs, that are highly glucose-sensitive ß-cells, on the overall network dynamics, observing different modulation depending on the timescale of the dynamics. By properly taking into account the role of cells heterogeneity, recently emerged, our model effectively describes the effect of hubs on the synchronization of the islet response and the correlation of ß-cells activity.

A possible desensitized state conformation of the human α7 nicotinic receptor: A molecular dynamics study.

The determination of the conformational states corresponding to diverse functional roles of ligand gated ion channels is subject of intense investigation with various techniques, from X-rays structure determination to electrophysiology and computational modeling. Even with a certain number of structures becoming recently available, only few major structural features distinguishing conductive open channel from the non conductive resting protein have been highlighted, while high-resolution details are still missing. The characterization of the desensitized conformation(s) is even more complex, and only few specific characteristics have been identified. Furthermore, experimental data provide conflicting information for different ion channels, adding further complexity to the topic. Desensitization is defined as the transition of the agonist-bound open channel into an ion channel configuration inactive even in the presence of agonists. In this work, we analyze a conformation corresponding to a non conductive state obtained via molecular dynamics simulations of a homology model of the human α7 nicotinic receptor complexed with agonists. We highlight some characteristics that could associate it to a desensitized state. The obtained structure is assessed against experimental data for other ligand gated ion channels that have been putatively associated to active, inactive and desensitized conditions.

Strongly bound excitons in anatase TiO2 single crystals and nanoparticles.

Anatase TiO2 is among the most studied materials for light-energy conversion applications, but the nature of its fundamental charge excitations is still unknown. Yet it is crucial to establish whether light absorption creates uncorrelated electron-hole pairs or bound excitons and, in the latter case, to determine their character. Here, by combining steady-state angle-resolved photoemission spectroscopy and spectroscopic ellipsometry with state-of-the-art ab initio calculations, we demonstrate that the direct optical gap of single crystals is dominated by a strongly bound exciton rising over the continuum of indirect interband transitions. This exciton possesses an intermediate character between the Wannier-Mott and Frenkel regimes and displays a peculiar two-dimensional wavefunction in the three-dimensional lattice. The nature of the higher-energy excitations is also identified. The universal validity of our results is confirmed up to room temperature by observing the same elementary excitations in defect-rich samples (doped single crystals and nanoparticles) via ultrafast two-dimensional deep-ultraviolet spectroscopy.Here the authors combine steady-state angle-resolved photoemission spectroscopy, ellipsometry and ultrafast two-dimensional ultraviolet spectroscopy to examine the role of many-body correlations in anatase TiO2, revealing the existence of strongly bound excitons in single crystals and nanoparticles.

Teens with heavy prenatal cocaine exposure respond to experimental social provocation with escape not aggression.

Preclinical data show that, compared to no exposure, prenatal cocaine exposure (PCE) has age-dependent effects on social interaction and aggression. The aim of this clinical study was to determine how heavy/persistent PCE--after controlling for other prenatal drug exposures, sex and postnatal factors--predicts behavioral sensitivity to provocation (i.e., reactive aggression) using a well-validated human laboratory model of aggression. African American teens (mean=14.2 years old) with histories of heavy/persistent PCE (maternal cocaine use ≥ 2 times/week during pregnancy, or positive maternal or infant urine/meconium test at delivery; n=86) or none/some exposure (NON: maternal cocaine use < 2 times/week during pregnancy; n=330) completed the Point Subtraction Aggression Paradigm. In this task, teens competed in a computer game against a fictitious opponent. There were three possible responses: (a) earn points, to exchange for money later; or (b) "aggress" against the fictitious opponent by subtracting their points; or (c) escape temporarily from point subtraction perpetrated by the fictitious opponent. The PCE group responded significantly more frequently on the escape option than the NON group, but did not differ in aggressive or money-earning responses. These data indicate that PCE-teens provoked with a social stressor exhibit a behavioral preference for escape (negative reinforcement) than for aggressive (retaliatory) or appetitive (point- or money-reinforced) responses. These findings are consistent with preclinical data showing that social provocation of adolescent or young adult offspring after PCE is associated with greater escape behavior, inferring greater submission, social withdrawal, or anxiety, as opposed to aggressive behavior.

Optical response of the copper surface to carbon monoxide deposition.

The optical response of the Cu surface upon CO deposition is investigated from the clean Cu(110) to the reconstructed CO/Cu(110)-p(2x1) geometry through ab initio electronic structure calculations. We ascribe the relevant structures in the calculated reflectance anisotropy spectrum of the reconstructed phase to the persistence of surface states transitions. These are excited by light polarized along the direction perpendicular to the one found at the clean surface. We devise a simple model for the evolution of the optical response in the adsorption process, identifying three different regimes. The impurity regime, at very low coverages, is characterized by a critical coverage that enhances the actual one by a factor of approximately 30, close to the value estimated experimentally.

Severe dysosmia is specifically associated with Alzheimer-like memory deficits in nondemented elderly retirees.

OBJECTIVES: To determine whether or not (1) impaired olfactory function is associated with impaired memory on neuropsychological testing in healthy retirees, and if so then (2) whether memory impairment is most consistent with a mesiotemporal rather than frontal system disorder. METHODS: 173 independent residents of a continuing care retirement community were studied. Subjects completed the University of Pennsylvania Smell Identification Test (UPSIT) and a battery of both general and specific cognitive measures that included the Mini-Mental State Examination (MMSE) and the Executive Interview (EXIT25). Subjects were examined twice over 3 years. RESULTS: UPSIT performance was normal in 21% and in the 'anosmic' range in 25% of subjects. Anosmic UPSIT performance was associated with significantly worse performance on all cognitive tests. However, only short-term verbal memory was independently associated with UPSIT-defined anosmia. This association remained significant after adjusting for the other cognitive and sociodemographic variables. The memory deficits of anosmic subjects were qualitatively consistent with a cortical type (type 1) dementing illness such as Alzheimer's disease (AD). Over time, UPSIT-defined 'anosmic' cases suffered significantly greater declines on both the MMSE and the EXIT25, independently of baseline age, gender and MMSE score. CONCLUSIONS: Impaired odor identification in individuals without overt dementia is associated with an AD-like memory impairment and an increased rate of cognitive decline. The comorbid association of these deficits is consistent with the known hierarchical spread of preclinical AD pathology and may be a specific indicator of future clinical AD dementia.

Correlates of disability among elderly retirees with "subclinical" cognitive impairment.

BACKGROUND: We assessed the effects of impaired Executive Control Function (ECF) on Instrumental Activities of Daily Living (IADL) and level of care among noninstitutionalized elderly retirees with "subclinical" cognitive impairment. METHODS: Subjects (N = 561; age 78.2 +/- 5.0 years) were residents of a single, 1,500 bed, continuing care retirement community. Subjects were examined for cognitive impairment using the Executive Interview (EXIT25), Mini-Mental State Examination (MMSE), and an executive clock-drawing task (CLOX). The CLOX is divided into executively sensitive (CLOX 1) and simple constructional (CLOX2) subtests. RESULTS: Residents in congregate high-rises (n = 301) differed significantly from those in independent-living apartments (n = 260) with respect to age, gender, percent living alone, EXIT25, CLOX1, MMSE, and CLOX2 scores (all p < .03). Only differences in ECF measures persisted after adjusting for age and living alone (p < .004). The EXIT25 (p < .006) and CLOX2 (p = .02) were associated with the use of prostheses. The differences in EXIT25 scores persisted after adjusting for level and living alone (p = .01). All instruments distinguished residents with impairment in IADLs. However, only CLOX2 (p < .001), EXIT25 (p < .001), and age (p < .001) made significant independent contributions. CONCLUSIONS: ECF has statistically significant effects on level of care and IADL impairment, even among noninstitutionalized retirees. This emergent disability is not well detected by traditional global cognitive measures. Evaluation and treatment may be delayed unless ECF measures are employed.

Clock drawing is sensitive to executive control: a comparison of six methods.

We examined six clock-drawing task (CDT) scoring systems relative to the Executive Interview (EXIT25, a measure of Executive Control Function [ECF]) and the Mini-Mental State Exam (MMSE). Subjects included n = 33 National Institute of Neurological, Communicative Disorders, and Stroke "probable" Alzheimer's disease (AD) cases and n = 52 independent living controls. AD cases and controls differed on the EXIT25, MMSE, and all CDTs. All CDTs were significantly correlated with the EXIT25 (ranging from r = .56 to r = .78). These associations generally persisted after adjusting for Age, Education, and MMSE scores. In backwards stepwise linear multivariate regression models, only CLOX: An Executive Clock-Drawing Task scores contribute significantly to EXIT25 scores (R2 = .68) and MMSE scores (R2 = .72). Clock drawing draws upon both executive and general cognitive resources. CLOX explains incrementally more variance in ECF than other CDTs.

Breastfeeding effects on intelligence quotient in 4- and 11-year-old children.

OBJECTIVE: A study of preterm children found an IQ advantage among those who were breastfed as infants after controlling for maternal social class and educational status. However, this advantage needs to be examined in light of other maternal characteristics, such as maternal IQ and parenting skills, which were not measured in that study and which have been found to be related to breastfeeding. METHODOLOGY: IQ was assessed in 323 children at 4 years of age on the McCarthy Scales of Children's Abilities and the Peabody Picture Vocabulary Test-Revised and in 280 children on the Wechsler Intelligence Test for Children-Revised at 11 years of age. RESULTS: Children who were breastfed in infancy had significantly higher scores on IQ tests at both ages, even after adjusting for social class and education, confirming the earlier findings and extending them to a predominantly full-term sample. However, the effect of breastfeeding was no longer significant after adjusting for maternal IQ assessed on the Peabody Picture Vocabulary Test-Revised and for parenting skills assessed on the Home Observation for Measurement of the Environment. Significant relations between breastfeeding and Woodcock Reading Achievement scores at 11 years were also reduced to nonsignificant levels after the inclusion of maternal IQ and the Home Observation for Measurement of the Environment. CONCLUSIONS: These findings suggest that the observed advantage of breastfeeding on IQ is related to genetic and socioenvironmental factors rather than to the nutritional benefits of breastfeeding on neurodevelopment. They should not be interpreted as detracting from the medical benefits associated with breastfeeding.

D2S, D2L, D3, and D4 dopamine receptors couple to a voltage-dependent potassium current in N18TG2 x mesencephalon hybrid cell (MES-23.5) via distinct G proteins.

We utilized the approach of stably expressing different dopamine (DA) receptors into identified cell lines in an attempt to better understand the coupling of these receptors to membrane ion channels via second messenger systems. Recently, we examined the N18TG2 x mesencephalon (MES-23.5) cell line that is phenotypically similar to mesencephalic dopamine-containing neurons. Whole-cell voltage-clamp methods were used to investigate a voltage-dependent K+ current present in these cells. Untransfected MES-23.5 cells displayed a voltage-dependent slow-onset, slowly inactivating outward current which was not altered by bath application of either the D2 DA receptor agonist quinpirole (QUIN; 10-100 microM) or the D1 DA receptor agonist SKF38393, indicating that these cells were devoid of DA receptors. The K+ current studied was activated upon depolarization from a holding potential of -60 mV to a level more positive than -20 mV and was observed to be sensitive to bath application of tetraethylammonium. When MES-23.5 cells were transfected to stably express the D2S, D2L, D3, and D4 receptors, the same current was observed. In cells expressing D2L, D2S, and D3 receptors, application of the DA receptor agonists QUIN (1-80 microM), 7-hydroxy-dipropylaminoteralin (7-OH-DPAT, 1-80 microM), and dopamine (DA, 1-80 microM), increased the peak outward current by 35-40%. In marked contrast, cells stably expressing the D4 receptor demonstrated a significant DA agonist-induced reduction of the peak K+ current by 40%. For all four receptor subtypes, the D2-like receptor antagonist sulpiride (SUL 5 microM), when coapplied with QUIN (10 microM), totally abolished the change in K+ current normally observed, while coapplication of the D1-like receptor antagonist SCH23390 was without effect. The modulation of K+ current by D2L, D3, and D4 receptor stimulation was prevented by pretreatment of the cells with pertussis toxin (PTX, 500 ng/ml for 4 h). In addition, the intracellular application of a polyclonal antibody which specifically recognizes Goalpha completely blocked the ability of D2L, D3, and D4 receptors to modulate outward K+ currents. In contrast, the intracellular application of an antibody directed against Goalpha was without effect, whereas intracellular application of an antibody recognizing Gsalpha abolished the ability of the D2S receptor to enhance K+ current. These findings demonstrate that different members of the D2 DA receptor family may couple in a given cell to a common effector in dramatically different ways.

Effects of prenatal alcohol and cocaine exposure on infant cortisol levels.

Elevated corticosterone levels to stress have been found in adult rats exposed prenatally to alcohol, but little is known about the effects of prenatal alcohol exposure on the cortisol response in humans. To date, one study has found that crack/cocaine was related to depressed newborn cortisol levels following a heel prick. In the present study saliva samples were obtained before and after a blood draw from 83 inner-city African American 13-month-old infants exposed prenatally to alcohol, cocaine, and other illicit drugs. Post-blood draw cortisol levels did not differ from basal levels in many of the infants, confirming recent studies indicating adaptation of the adrenocortical response to this type of stress at this age. Maternal depression and emergence of teeth were positively related to cortisol levels. Alcohol exposure was related to elevated basal levels, cocaine to lower basal levels. As predicted from animal findings, heavy alcohol exposure was related to elevated poststress cortisol levels.

New evidence for neurobehavioral effects of in utero cocaine exposure.

Most studies of prenatal cocaine exposure have found gestational age or intrauterine growth deficits but few, if any, cognitive effects. In a large, well-controlled study we detected cognitive deficits in relation to heavy cocaine exposure. These findings demonstrate that prenatal exposure to cocaine at sufficiently high doses early in pregnancy has the potential to produce cognitive changes in infants and that more focused, narrow-band tests may be necessary to detect these subtle neurobehavioral effects. A total of 464 inner-city, black infants whose mothers were recruited prenatally on the basis of pregnancy alcohol and cocaine use were tested at 6.5, 12, and 13 months of age. Standard analyses, based on presence or absence of cocaine use during pregnancy, confirmed effects on gestational age but failed to detect cognitive effects. A new approach to identifying heavy users found that heavy exposure early in pregnancy was related to faster responsiveness on an infant visual expectancy test but to poorer recognition memory and information processing, deficits consistent with prior human and animal findings. These persistent neurobehavioral effects of heavy prenatal cocaine exposure appear to be direct effects of exposure and independent of effects on gestational age.

D2L, D2S, and D3 dopamine receptors stably transfected into NG108-15 cells couple to a voltage-dependent potassium current via distinct G protein mechanisms.

The D2-like dopamine (DA) receptor family has continued to expand and now includes the D2-short (D2S) and D2-long (D2L) receptor isoforms and the D3 and D4 receptors. The D2 receptor isoforms differ in length by 29 amino acids within the third cytoplasmic loop, a region of the receptor believed to be important for G protein coupling. This observation has led to the hypothesis that the two isoforms of the D2 receptor may utilize different signal transduction pathways when present in the same cell. The D2 and D3 receptors, although mostly different, show some common amino acid sequences within the third cytoplasmic loop. Thus, it is possible that the D2 and D3 receptors may employ similar signal transduction pathways. To test these hypotheses directly, NG108-15 neuroblastoma-glioma hybrid cells were stably transfected to express either the D2S, D2L, or D3 DA receptors. All transfected but not untransfected NG108-15 cells demonstrated a dose-dependent reduction in the peak whole-cell potassium (K+) current in response to receptor activation by DA or the DA receptor agonists quinpirole (QUIN) and apomorphine (APO). The modulation of K+ current by D2S receptor stimulation was prevented by pretreatment of the cells with cholera toxin (20 micrograms/ml for 18 h), whereas pertussis toxin pretreatment (500 ng/ml for 4 h) completely blocked the effects of D2L and D3 receptor activation. These observations suggest that the signal transduction mechanisms involved in coupling the two isoforms of the D2 receptor to the K+ current are different, whereas the D2L and D3 receptor coupling mechanisms may be similar. In direct support of this hypothesis, it was observed that the intracellular application of a polyclonal antibody that is specific for the GO alpha subunit completely blocked the ability of D2L and D3 receptors to modulate outward K+ currents. In contrast, the D2S-mediated modulation of K+ currents was blocked by intracellular application of an antibody recognizing GS alpha but not GO alpha. These findings demonstrate that D2S and D2L receptors are able to couple to a common effector in a cell via two G protein pathways.

Function and medical comorbidity in south Texas nursing home residents: variations by ethnic group.

OBJECTIVE: To evaluate differences in functional status and burdens of medical conditions in Mexican American and non-Hispanic white nursing home residents. DESIGN AND SETTING: Cross-sectional survey of 17 nursing homes in south Texas. PARTICIPANTS: A total of 617 older nursing home residents, of whom 366 were Mexican American and 251 were non-Hispanic white. MEASURES: Activities of Daily Living (ADL) status abstracted from standard nurses notes and Burden of Disease abstracted from medical records. RESULTS: Mexican American residents had greater numbers of ADL dependencies and poorer overall ADL scores than non-Hispanic white residents. This poor functioning was not explained by age, gender, or marital or educational status. The average number of medical conditions was greater, and specific conditions, such as cerebrovascular disease, recent acute infections, diabetes, hypertension, and anemia, were more common in Mexican American residents compared with non-Hispanic white residents. In models relating function with medical conditions and ethnic group, ADL scores and dependencies were significantly related to bowel and bladder incontinence, cerebrovascular disease, dementia, recent infections, and skin decubiti, but not to ethnic group. CONCLUSION: Mexican American nursing home residents are more functionally dependent than non-Hispanic white residents. The difference in function is explained by a greater burden of medical conditions in the Mexican American residents.

Discovery of a brain promoter from the human transferrin gene and its utilization for development of transgenic mice that express human apolipoprotein E alleles.

Transgenic mice carrying heterologous genes directed by a 670-bp segment of the regulatory sequence from the human transferrin (TF) gene demonstrated high expression in brain. Mice carrying the chimeric 0.67kbTF-CAT gene expressed TF-CAT in neurons and glial cells of the nucleus basalis, the cerebrum, corpus callosum, cerebellum, and hippocampus. In brains from two independent TF-CAT transgenic founder lines, copy number of TF-CAT mRNA exceeded the number of mRNA transcripts encoding either mouse endogenous transferrin or mouse endogenous amyloid precursor protein. In two transgenic founder lines, the chloramphenicol acetyltransferase (CAT) protein synthesized from the TF-CAT mRNA was estimated to be 0.10-0.15% of the total soluble proteins of the brain. High expression observed in brain indicates that the 0.67kbTF promoter is a promising director of brain expression of heterologous genes. Therefore, the promoter has been used to express the three common human apolipoprotein E (apoE) alleles in transgenic mouse brains. The apoE alleles have been implicated in the expression of Alzheimer disease, and the human apoE isoforms are reported to interact with different affinities to the brain beta-amyloid and tau protein in vitro. Results of this study demonstrate high expression and production of human apoE proteins in transgenic mouse brains. The model may be used to characterize the interaction of human apoE isoforms with other brain proteins and provide information helpful in designing therapeutic strategies for Alzheimer disease.

Repeated stimulation of dopamine D2-like receptors: reduced responsiveness of nigrostriatal and mesoaccumbens dopamine neurons to quinpirole.

Extracellular recording techniques were used to study antidromically activated nigrostriatal (NSDA) and mesoaccumbens (MADA) dopamine neurons in chloral hydrate-anesthetized rats. Repeated 14-day i.p. treatment with the dopamine D2-like receptor agonists, quinpirole (2 mg/kg/day) or EMD 23448 (2.6 mg/kg/day), resulted in a significant decrease in the average potency and efficacy of i.v. quinpirole (cumulative doses administered on day 15) to inhibit the spontaneous activity of NSDA neurons relative to vehicle controls. Repeated 14-day quinpirole treatment caused a significantly greater decrease in the sensitivity of MADA neurons to i.v. quinpirole challenges than NSDA neurons. When the effects on NSDA neurons were examined after a shorter treatment period, the decrease in the average potency and efficacy of i.v. quinpirole appeared to occur after only 2 days of i.p. quinpirole treatment (2 mg/kg/day). Iontophoretic studies, however, indicated that the average dopamine sensitivity of somatodendritic dopamine autoreceptors on MADA neurons, but not NSDA neurons, was significantly lower relative to controls after 14-day quinpirole treatment (2 mg/kg/day). These results suggest that this quinpirole treatment regimen can differentially affect the average sensitivity of somatodendritic dopamine autoreceptors on MADA and NSDA neurons. The somatodendritic autoreceptors on MADA neurons appear to be more sensitive to the effects of repeated 14-day quinpirole treatment than those on NSDA neurons.

Representations of self across the life span.

This research extends a cognitive-developmental approach to examining age differences in self-representation from adolescence to mature adulthood and later life. The authors suggest that mature adults move from representations of self that are relatively poorly differentiated from others or social conventions to ones that involve emphasis on process, context, and individuality. Participants (n men = 73, n women = 76), ranging in age from 11 to 85 years, provided spontaneous accounts of their self-representations and responded to measures assessing cognitive and emotional functioning and broad dimensions of personality. On average, self-representation scores peaked in middle-aged adults and were lowest in the preadolescent and older adult age groups. Level of self-representation was related to cognitive and personality variables, but there was some evidence that the pattern of correlates shifted from younger (ages 15-45) to older (ages 46-85) age segments.

The effects of chronic amphetamine treatment on prenatal ethanol-induced changes in dopamine receptor function: electrophysiological findings.

The sensitivity of dopamine (DA) receptors in the mesoaccumbens DA system was investigated with extracellular recording and microiontophoresis techniques in adult rats that received prenatal ethanol exposure and chronic postnatal amphetamine treatment. Pregnant rats were fed with a liquid diet containing 0 or 35% ethanol-derived calories from gestation day 6 to 20. An ad libitum group received laboratory chow and water. Offspring were injected with amphetamine (2 mg/kg/day s.c.) or saline from postnatal day 22 to 10- to 12-months of age. Electrophysiological recording procedures were performed 16 to 24 hr after the last amphetamine injection. A supersensitivity of somatodendritic DA autoreceptors in the ventral tegmental area was observed in animals exposed prenatally to ethanol. This prenatal ethanol exposure-induced supersensitivity was not observed after postnatal amphetamine treatment. In control animals, postnatal amphetamine treatment did not affect the sensitivity of somatodendritic DA autoreceptors. The sensitivity of D-1 DA receptors in the nucleus accumbens was reduced by prenatal ethanol exposure. Postnatal amphetamine treatment reduced D-1 DA receptor sensitivity in control animals, but not in animals exposed prenatally to ethanol. Neither prenatal ethanol treatment nor postnatal amphetamine treatment altered the sensitivity of D-2 DA receptors in the nucleus accumbens. There were no differences between the ad libitum and 0% ethanol-derived calorie groups, indicating undernutrition did not affect DA receptor function. These results show that prenatal ethanol exposure altered DA receptor function in the mesoaccumbens DA system in adult animals. Furthermore, postnatal amphetamine treatment was able to eliminate the supersensitivity of somatodendritic DA autoreceptors in prenatal ethanol-exposed animals.