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Medical
and Health Sciences
Abstracts:
9- THC Chronic Exposure in Adolescent Rats: Effects on Cocaine Conditioned
Place Preference. YU FEN HWANG (State University
of New York at Stony Brook, Stony Brook, NY, 11794) PETER THANOS (Brookhaven
National Laboratory, Upton, NY, 11973)
Epidemiological studies have
shown that use of marijuana during early adolescence is associated with a higher
risk for other substance abuse disorders. The extent to which this represents
neuroadaptation responses secondary to 9-THC (delta-9-tetrahydrocannbinol, main
psychoactive ingredient of marijuana), exposure versus genetic vulnerability
that underlies high risk with experimentation with drugs in general is unclear.
In this study we investigated if chronic exposure of 9- THC during the adolescence
period in the rat affected the reinforcing responses to cocaine (assessed with
condition place preference). Male Sprague Dawley rats (age 3-4 weeks) were administered
daily with either 9- THC (0.75 and 2 mg/kg) or saline for 21 days. The day after
the last injection, we started the Conditioned Place Preference (CPP) segment
of the study: [day 1: preconditioning; days 2-9 conditioning phase {cocaine (5
and 10mg/kg) and saline on alternate days}; day 10: test day]. The results showed
that 9- THC pretreatment rats resulted in decreased CPP for cocaine (5mg/kg
or 10mg/kg) On the other hand there were no differences between the group on
cocaine induced dose-dependent increases in locomotor activity. These results
provide evidence of that exposure to THC during the adolescent period changes
the sensitivity to cocaine-induced conditioning. The extent to which this generalizes
to conditioning to other drug but also non-drug reinforcers requires further
investigation.
A Personality Profile of Cocaine Addicted Individuals. CATHERINE
URBAN (State University of New York at Geneseo, Geneseo, NY, 14454) DR.
RITA GOLDSTEIN (Brookhaven National Laboratory, Upton, NY, 11973)
Impairments in higher executive
function, such as inhibitory control and decision-making, characterize cocaine
addicted individuals; however, the contribution to these cognitive functions
of personality traits is not well established. The current study therefore examined
differences in personality traits between healthy control subjects and cocaine
addicted individuals and the extent to which these differences may be affected
by recent cocaine use. Sixty-six cocaine addicted individuals [20 testing negative
(CUD-) and 46 testing positive (CUD+) for cocaine in urine, indicative of drug
use within 72-hours] and 66 gender-, age-, and education-matched healthy control
subjects were administered the Multidimensional Personality Questionnaire (MPQ)
as part of a larger neuropsychological battery. The MPQ measures three trait
super-factors, each comprised of 3-4 lower order subscales; differences between
the groups were examined with four separate MANOVAs. Results revealed significant
group differences on all three super-factors such that cocaine abuse was associated
with lower positive emotionality [F(2, 131) = 3.8, p < .0001],
higher negative emotionality [F(2,131)
= 5.0, p < .0001], and
decreased constraint [F(2,
131) = 3.1, p < .001]. These differences were driven by lower social closeness
especially in the cocaine positive subgroup [Mean±SEM=12.3±0.6
for CUD+ vs. 14.0±0.8
for CUD- vs. 15.7±0.5 for controls], higher aggression especially in the cocaine
negative subgroup [Mean±SEM=5.7±0.5 for CUD+ vs. 6.7±1.0 for CUD- vs. 3.4±0.4 for controls],
and lower self-control in the cocaine positive subgroup [Mean±SEM=14.1±0.7
for CUD+ vs. 16.0±1.1
for CUD- vs. 17.7±0.5 for controls]. These effects remained significant after
controlling (with ANCOVAs) for state depression and other demographic variables
that differed between the groups. These results suggest significant differences
between healthy control subjects and individuals addicted to cocaine in trait
measures of personality. Together, the higher negative emotionality (aggression),
lower positive emotionality (social closeness) and lower self-control may predispose
the addicted individuals to relapse and compulsive drug use, especially under
socially stressful situations. The underlying cognitive and neurobiological substrates
remain to be elucidated.
A Review of Empirical Methods for the Derivation of Parameters in a Theoretical
Model of Matrix Metalloproteinase 2 & 9 Proteolysis of Collagen
Type IV. ELIZABETH O'QUINN O'QUINN (Wofford College, Spartanburg,
SC, 29303) KARA KRUSE (Oak Ridge National Laboratory, Oak Ridge,
TN, 37831)
Cardiovascular disease
is the leading cause of death in first world countries. An imbalance of matrix
degrading enzymes and structural proteins within the extracellular matrix of
an arterial wall is a critical factor in cardiovascular disease processes.
An increase in matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9
(MMP-9), as part of the inflammatory process, results in degradation of collagen
type IV influencing the migration and proliferation of vascular smooth muscle
cells; this can lead to further narrowing of a diseased artery. Kinetic modeling
of proteolysis is an approach which can be used to understand complex systems
by describing enzymatic mechanisms, cellular processes, and the system’s behavior
quantitatively. In this research project, a computational model of the biochemical
pathways involved in activation and inhibition of MMP-2 and MMP-9 proteolysis
of collagen type IV is being developed from empirical data and published data.
Separate and integrated models of MMP-2 and MMP-9 pathways have been implemented
within JSim, a software application developed by the University of Washington.
In addition to the enzyme model a cellular migration model is also being developed
for the simulation of VSMC migration and will be explored further. The utilization
of reverse-phase high pressure liquid chromatography (HPLC) methods for obtaining
quantitative reaction rate parameters are being explored for the estimation
of parameters not previously published in the literature. By pairing HPLC separation
with spectrometry techniques, protein and peptide identification and quantification
are possible. Experimental protocols for the measurement of the enzymatic activity
of MMP-2 and MMP-9 proteolysis of collagen type IV are being developed to obtain
empirical data. These experimental results are then analyzed to derive the
rate parameters needed in the computational model. The use of HPLC methods
to analyze the enzymatic activity and cellular activity provides parameters
which cannot be obtained through literature. This research is in collaboration
with the Vascular Research Laboratory at the University of Tennessee Medical
Center in Knoxville.
Analysis of the Biological Effects of Aspirated Carbon Nanohorn Particles in
Mice using Scanning Near-Field Ultrasound Holography. KATHERINE
VENMAR (Denison University, Granville, OH, 43023) THOMAS THUNDAT
(Oak Ridge National Laboratory, Oak Ridge, TN, 37831)
Engineered nanomaterials,
because of their enhanced physicochemical properties compared to their
bulk form, are finding an increasingly important role in many potential
commercial applications. However, the health effects of nanomaterials
are not well understood or thoroughly investigated. Therefore, more studies
are needed to examine different types of nanomaterials and the biological
responses they invoke. The purpose of this research was to examine the
effects of aspirating single-walled carbon nanohorns (SWCNHs) in vivo
using mice. Bronchoalveolar lavage (BAL) and blood samples were collected
from two experimental groups, the nanohorn exposed, and the control mice.
Three mice from both groups were sacrificed 24 hours and 7 days after
aspiration. Gross examination of the number of macrophages versus activated
macrophages in BAL samples from the exposed and the control mice suggested
a possible pro-inflammatory response to the carbon nanohorns. Employing
a unique detection technique, Scanning Near-Field Ultrasound Holography,
carbon nanohorns were discovered bound to cell membranes, inside cells,
and near cells in both the red blood cells and BAL sample cells. The
positioning of carbon nanohorns inside the cells not bound to a membrane
suggests that they entered the cell through a process other than phagocytosis.
Furthermore, the red blood cells (RBC) in all the exposed blood samples
exhibited a distorted phenotype. Such distortions could possibly lead
to various pulmonary diseases. From their ability to permeate membranes,
cause pro-inflammatory responses, and distort the phenotype of red blood
cells, it can be concluded that carbon nanohorns may pose a biological
threat.
Biodistribution and Metabolism Studies of 14C-Ethanol in Mice. MEL PILAR
ESPAILLAT (Farmingdale State College, Farmingdale, ny, 11735) ANDREW
GIFFORD (Brookhaven National Laboratory, Upton, NY, 11973)
Ethanol acts as a central
nervous system depressant; it is rapidly absorbed by the body and approximately
90% is metabolized, almost entirely in the liver. The intermediate metabolites
of ethanol are believed to be involved in its toxicological properties. The mechanism
of ethanol toxicity and its interaction with tissue have not been well understood
because of lack of fundamental knowledge about both the pharmacokinetics of ethanol
and its metabolism in the human body. This work seeks to study the rate of metabolism
of ethanol in different tissues and provide preliminary data for future PET studies
to directly measure the distribution and pharmacokinetics of ethanol in human
subjects. Mice
were injected with approximately
2.0x10-¬¬3 microcuries of 14C-Ethanol and then were sacrificed at 5, 15, and
60 minute intervals after injection. Plasma and various organs were collected
and assayed for carbon-14 under acidic and basic conditions. In the present study
a high concentrations non-volatile material was found in the liver, consistent
with the role of this organ in metabolizing ethanol. However it is notable that
other organs were also found to accumulate both ethanol and non-volatile metabolites
although it is not possible to say whether this represents accumulation of circulating
radiolabeled metabolites from liver metabolism of ethanol or is due to local
metabolism of the radiolabeled ethanol. This work is a small portion of a much
larger project being researched to investigate the distribution and pharmacokinetics
of ethanol in human subjects.
Chronic THC Exposure: Effects on Sucrose Conditioned Place Preference in Adolescent
Rats. ANNA VERDE (State University of New York at Stony
Brook, Stony Brook, NY, 11790) PANAYOTIS (PETER) THANOS (Brookhaven
National Laboratory, Upton, NY, 11973)
The psychoactive constituent
in marijuana 9-tetrahydrocannabinol (THC) pharmacologically activates the mesolimbic
reward pathways. Similarly highly palatable foods also activate the reward circuitry
of the brain. Specifically, administration of THC has been shown to influence
the intake of sweet foods. The effect of THC in adolescence
hasn’t been looked at yet. Adolescence may be characterized as a period when
a significant amount of neurobiological and development changes occur. Drug abuse
during an early neurodevelopmental period may impact the reward potential and
consumption of foods or drugs later in life. Therefore, the goal of the present
study was to examine the effect of chronic THC administration during adolescence
on the reward potential of sucrose using a conditioned place preference (CPP)
paradigm. During the exposure period, 4 week old male Sprague Dawley rats were
divided into 3 groups to receive a daily i.p. injection for 3 weeks of either:
1) vehicle (saline) 2) low dose THC (0.75 mg/kg) 3) high dose THC (2 mg/kg).
Next, all rats started CPP after the last day of treatment. The CPP timeline
encompassed the following: Day 1: Habituation, Days 2-9: Conditioning Phase (10
sucrose pellets on even days and no sucrose on odd days), and Day 10: Test Day.
These findings will help gain insight on the impact of chronic THC exposure during
a neurodevelopmental period on the subsequent reward potential of natural rewards.
Finally these sucrose CPP results will be compared to CPP results to drugs in
similar THC pretreated rats (Hwang et al. 2006).
Conflict Resolution is Impaired in Currently Withdrawn Cocaine Addicted Individuals. BAABA BLANKSON (State University of New York at Stony
Brook, Stony Brook, NY, 11790) RITA GOLDSTEIN (Brookhaven National Laboratory,
Upton, NY, 11973)
Individuals with cocaine
use disorders (iwCUD=CUD) experience neuropsychological impairments that encompass
attention and executive function deficits, possibly due to structural and functional
changes in prefrontal cortical brain regions. The Attention Network Test (ANT),
a computerized reaction time task, has been developed to measure three of these
attention and executive function networks: alerting (associated with right hemisphere
frontal and parietal regions), orienting (associated with subcortical regions),
and executive control (associated with the prefrontal cortex, particularly the
anterior cingulate cortex). Our study used the ANT to examine 1) differences
between CUD and healthy control subjects in attention and executive function;
and 2) the effect of urine status (positive or negative for cocaine use within
72 hours) on attention and executive function in the CUD group. The ANT was administered
as part of a larger neuropsychological battery to 84 individuals with current
CUD [55 positive (CUD+), 29 negative (CUD-)] and 75 healthy control subjects
matched on gender, age, and years of education, but not on race, general intellectual
functioning, socioeconomic status, state depression and history of cigarette
smoking. Results of 3 separate ANOVAs revealed a significant group difference
in executive control [[F (2,
156) = 5.8, p < 0.004;
pairwise comparisons: controls faster than both cocaine subgroups, an effect
that reached significance
for CUD-, p < .004, M±SD =
127.5±47.8 for controls vs. 149.4±64.5 for CUD+ vs. 169.8±76.5 for CUD-], but
not in orienting [F (2,
158) = 1.0, p > 0.4]
or alerting [F (2,
157)
= 0.2, p > 0.8]. The executive control effect remained significant after controlling
(with ANCOVA) for general intellectual functioning, the only non-matched variable
that correlated with executive control (p < 0.03).
The current results suggest that the ability to resolve a cognitive conflict
may be uniquely impaired in CUD and expressed primarily during acute withdrawal.
Possible explanations may encompass the self-medicating hypothesis (i.e., where
cocaine use ameliorates an underlying cognitive deficit in CUD) or the effects
of acute withdrawal symptoms on neuropsychological function. The effects of longer
abstinence periods and other drug use variables on ANT conflict in CUD remain
to be established. Future studies need to also explore the role of the anterior
cingulate cortex in underlying this cognitive conflict resolution performance
deficit in drug addiction.
Determining the Optical Properties of Biological Tissue Samples Using an Integrating
Sphere Method. MARCUS ALLEGOOD (North Georgia
College & State University, Dahlonega, GA, 30553) JUSTIN S BABA
(Oak Ridge National Laboratory, Oak Ridge, TN, 37831)
Wavelength dependent light
interaction with biological tissue can be described using three parameters:
the scattering and absorption coefficients and the cosine of the average scattering
angle (g). To accurately determine these optical properties for different types
of tissue at specific wavelengths would be beneficial for a variety of different
biomedical applications. The goal of this project was to take a user defined
g-value and determine the remaining two parameters for a specified range of
wavelengths. In order to collect the needed data for all the wavelengths in
a timely and accurate manner, a fully automated computer program and process
was developed. Using a single integrating sphere method, scattered light intensity
inside the sphere was recorded via a spectrometer as either transmitted or
reflected light from the tissue sample. LabVIEW was used to write programs
to collect raw intensity data from the spectrometer, to convert the data into
a format for C code execution, and to compute the optical properties based
on the collected data. To make the process fully automated, the LabVIEW and
C code programs were linked together into one single program to allow data
to be passed between the two efficiently. The automated program was tested
using a tissue mimicking phantom and determination of the absorption and scattering
coefficients showed excellent agreement with theory. Future work and the final
phase of testing will entail examining actual biological tissue with known
optical properties to check for accuracy before proceeding to utilize the system
for its intended purpose. Ultimately, the data collection process and algorithms
developed through this effort will be applied to build models for light interaction
with biological
tissue samples.
Dose-Dependent Conditioned Place Preference Response and Locomotor Activity
in Methamphetamine-Treated Animals. REEMA
DALAL (New York University, New York, NY, 10003) DR. STEPHEN DEWEY
(Brookhaven National Laboratory, Upton, NY, 11973)
Previous behavioral studies
indicate that animals exposed to methamphetamine (METH) often associate and prefer
specific environments with their exposure, a behavior termed conditioned place
preference. The conditioned place preference (CPP) paradigm is a commonly used
technique to evaluate the positive or negative reinforcement values of different
drugs. These studies closely parallel behavioral responses reported by human
substance abusers. In the present study, the procedure involved several drug
pairings where the animals, adolescent male Sprague-Dawley rats, were exposed
to different doses of METH in distinct chambers containing well-defined tactile,
visual, and/or olfactory cues. Following randomization of the animals, the goal
was to expand previous studies, demonstrating that rats will express a CPP to
METH exposure by determining a dose-response. For these CPP studies, three groups
of animals were exposed to METH at concentrations of 2.5, 5.0, or 10.0 mg/kg.
Following twenty consecutive days of pairings (alternating with saline), animals
were given access to both chambers in a drug-free state (saline injection). The
amount of time spent in the METH-paired chamber was recorded, and was used as
an indicator of preference. Further, the locomotor activity was monitored each
day and the values obtained on the test day were compared against those obtained
for the pre-test. This analysis indicated a significant increase from low-dose
to high-dose administration, demonstrating that methamphetamine produced a significant,
dose-dependent CPP. Further studies using different concentrations are currently
ongoing in a continuing effort to better understand the locomotor and behavioral
effects
of METH.
Effect of stroke on regional distribution of aromatase and NMDA receptors in
the female rat brain. CANDACE GIRARD (Mount Holyoke
College, South Hadley, MA, 1075) ANAT BIEGON (Brookhaven National Laboratory,
Upton, NY, 11973)
The production of estrogens
from precursor androgens is catalyzed by the enzyme aromatase (AR), which under
normal circumstances, is restricted to specific neuronal locations. The N-methyl-D-aspartate
receptor (NMDAR) is one of the three major subtypes of glutamate receptors, which
under normal circumstances, plays a critical role in synaptic plasticity mechanisms.
Experimental data suggests that AR activity and NMDARs may be involved in neurodegenerative
processes following brain injury. Recent studies have indicated that brain insults
can induce brain AR expression and decrease brain NMDAR’s. However, the
long term effect of stroke w/respect to AR and NMDAR distribution has not been
investigated. Middle cerebral artery occlusion (MCAo) in female rats induced
stroke and was confirmed in the left hemisphere by MRI and histology. Stroke
rats were sacrificed 2 or 4 weeks post MCAo and brain tissue processed for autoradiography.
Five control brains (w/surgery; w/o stroke) and five experimental brains (w/surgery;
w/stroke) were used in these experiments. Using quantitative in vitro autoradiography,
AR distribution was investigated with [11C]-vorozole and NMDA distribution with
[3H]-MK-801. Regional densities of autoradiograms were quantified with standard
image analysis software. The [11C]-vorozole autoradiograms did not show specific
or increased binding in our stroke animals. This may be due to the low AR expression
in young females and/or a short-lived (<2wks) increase in aromatase. The [3H]-MK-801
autoradiograms showed similar regional binding density in control rats at 2 and
4 weeks. In stroke animals, binding density was decreased at 2wks after surgery
compared to controls as well as to stroke rats 4wks after surgery. These effects
were more pronounced in brain regions adjacent to the stroke. The most significant
increase in binding between 2 and 4wks after stroke was found to be in the frontal
cortex (~20%) and denate gyrus (~28%). Binding in the left caudate putamen was
significantly lower in experimental rats (~18%). These results, if upheld in
a larger series, suggest that hippocampal and cortical NMDAR reduction induced
by MCAo is transient, which is in line with the transient nature of many cognitive
symptoms often seen in the acute phase after a stroke. This also suggests that
damage to the caudate putamen may be sustained or irreversible, in line with
the long term nature of motor deficits among MCAo stroke patients.
Food Stimulation in a Rat Model of Obesity: Food Seeking Behavior as Assessed
by Nose-Poke Activity and Locomotor Responses in the Open-Field. LISA ROBISON (Colgate University, Hamilton, NY, 13346)
PETER THANOS (Brookhaven National Laboratory, Upton, NY, 11973)
Food intake is regulated
by factors
that modulate caloric requirements, as well
as food’s reinforcing properties. Apart from genetic factors, there are also
environmental factors that play a very important role in obesity, including society,
culture, stress, food palatability and food availability, as well as food-related
cues. Previously, we showed that exposure to an appetitive food stimulus had
significant effects on brain glucose in both humans (Wang et al. 2001) and rodents
(Michaelides et al 2006). In this study we examine the effect of genetically-induced
obesity and food restriction on seeking and learned behavior before and after
classical conditioning of a food (bacon) olfactory stimulus. Zucker Obese (Ob)
and Lean (Le) rats were divided into 4 groups: i) Ob ad lib fed, ii) Ob food
restricted (70% of ad-lib), iii) Le ad lib fed and iv) Le food restricted rats.
Leptin-receptor deficiency mixed with an unrestricted diet showed the greatest
significant amount of novelty-seeking.
Measurement of Methamphetamine-Induced Cue Response in Rodents using MicroPET. JOSEPH CARRION (The City College of New York, New
York, NY, 10033) DR. WYNNE SCHIFFER (Brookhaven National Laboratory,
Upton, NY, 11973)
Methamphetamine (Meth) has
seen an increase in use among eighth to twelfth graders. The effects of METH
are similar to amphetamine but even more potent and addictive to the central
nervous system. A behavioral model of craving with metabolic imaging using Micro
Positron Emission Tomography (MicroPET) was used to determine brain region activation
due to drug craving in response to specific stimuli. Conditioned Place Preference
(CPP) is an established method of assessing drug-paired environmental responses:
Cravings. Twelve male Sprague-Dawley rodents were injected with 5 mg/kg Meth
or saline and placed in distinct CPP chambers on alternating days. One day after
the 10th
Meth pairing, the CPP test was performed while the animal’s preference
was measured in a drug-free state. From these results, the preference score was
calculated as time spent in the saline-paired chamber minus time spent in the
Meth-paired chamber. Two days after the last Meth-pairing, animals received an
intraperitoneal (i.p.) injection of 18-Fluorodeoxyglucose (18-FDG), and were
placed in the Meth-paired chamber. After 45 minutes of FDG uptake, animals were
anesthetized and scanned in the MicroPET for 10 minutes. The animals were allowed
to rest for one day. Following the rest period, the animals were injected with
18FDG and placed in the saline-paired chamber. The same protocol was followed
for all twelve rodents. There was a significant preference for the METH paired
chamber (conditioned animals spent 461 sec in the METH paired chamber compared
to 260 sec in the saline paired chamber, p=0.002). Brain 18FDG data were analyzed
using Statistical Parametric Mapping (SPM) and Region of Interest (ROI) methodologies
correlated with a preference score. There were significant (p<0.01) 18FDG
increases in the primary somatosensory cortex, inferior colliculus, caudate putamen
and striatum. There were significant decreases in 18FDG uptake in the visual
cortex, retrosplenial cortex, and cingulate gyrus. There was a positive correlation
(p<0.03) between percent 18FDG uptake and preference score in the sensory
and memory regions of the brain. In these areas, FDG uptake increased with increasing
preference for the METH paired chamber. There were significant (p<0.01) negative
correlations in the piriform cortex, cerebellum and brainstem. In these areas,
FDG uptake increased with decreasing preference score. Here, we show that simple
exposure to a METH paired environment (without the choice) activates a network
of regions involved with spatial learning and recall.
Microbeam Radiation Therapy: Dose Distribution Calculations and Mathematical
Modeling. JENNIFER ONG (University of Illinois at Urbana-Champaign,
Urbana, IL, 61801) F. AVRAHAM DILMANIAN (Brookhaven National Laboratory,
Upton, NY, 11973)
Microbeam radiation therapy
(MRT) uses arrays of parallel, thin, synchrotron-generated x-ray beams, which,
when administered at high dosages, spare normal tissues while damaging tumor
tissues, thus exhibiting
a “preferential tumoricidal” effect. Implementing MRT in a new stereotactic
geometry (SMRT) of cylindrical pencil beams in circular arrays concentrates radiation
on the tumor by positioning it at the crossing of ten arrays while exposing surrounding
normal tissue merely to single arrays, thereby maximizing absorbed dose to the
tumor and minimizing damage to healthy tissues. In order to utilize SMRT geometry
in animal model experiments and ultimately on human carcinoma, determination
of the beam port angles, as well as an analysis of the spatial distribution of
the dose in exposed tissues, is needed. The irradiation port positions for the
new geometry were calculated using the computer program Matlab and then applied
to the rat brain tumor model experiments conducted at the National Synchrotron
Light Source (NSLS). The Monte Carlo code MCNP5 was then employed to simulate
the absorbed radiation dose profiles resulting from a 309-microbeam SMRT array
with radius 1.5cm in a spherical water phantom model of a human brain and tumor.
Superimposing a volumetric mesh tally of 150 µm x 150 µm x 1 cm voxels
on the phantom and using a photomultiplier in order to obtain absorbed dose from
1 billion incident photon events yielded relative errors <0.04% for the valley
dose, and showed that absorbed dose at the edge of a single beam decreases by
three orders of magnitude <450µm from each beam center. Furthermore,
the valley dose between beams is ~5% of the full, in-beam dose in the entire
1.5 cm-radius array at the center of the 7.5 cm-radius water sphere. These findings
confirm that microbeam dose dispersion in large subjects is small, and that the
valley dose remains low enough to allow normal-tissue repair stimulated by cells
surviving between beams. Future SMRT modeling should include the incorporation
of electron transport to account for energy deposition from photo- and Compton
electrons, the inclusion of more anatomically realistic brain/tumor geometry
and composition, and the calculation of dose profiles resulting from ten arrays
at SMRT angles.
Microbeam Radiation Therapy: Treating the 9LGS Rat Brain Tumor. NICOLLE LANIER (Suffolk County Community College,
Selden, NY, 11727) DR. AVRAHAM DILMANIAN (Brookhaven National Laboratory,
Upton, NY, 11973)
In conventional radiation
therapy the limitation in delivering the adequate dose to the tumor is the damage
to the normal surrounding tissue. Microbeam radiation therapy (MRT) is an experimental
method that uses arrays of parallel, thin (< 100 µm) planes of synchrotron-generated
x rays (called microbeams, or microplanar beams). It allows the normal tissue,
including the central nervous system (CNS), to recover from these irradiations
at doses many times higher than doses from the conventional beams that severely
damage the tissue. The effect has been attributed primarily to the quick recovery
of the microvasculature due to the survival of angiogenic cells between the microbeams.
Additionally, at very high doses, single-direction microbeam irradiation kills
the tumor in a single session without damaging the surrounding normal tissues,
an effect called "the preferential tumoricidal
effect of microbeams". However, our recent studies showed that the CNS also
tolerates beams as thick as 0.68 mm. We used an array of thick microbeams in
a method called "stereotactic microbeam radiation therapy (SMRT)" to
treat the rat brain tumor 9L gliosarcoma (9LGS). This method uses irradiations
from ten different angles, all crossing the tumor. The rats were irradiated with
SMRT at day 15-17 after inoculation (usually done at day 14). Following the irradiation
the rats were weighed and otherwise observed daily. The parameters registered
were a) if the rat was lethargic, b) if there is any sign of dry blood around
the rat’s eyes or nose (which are indications of intracranial pressure produced
by the tumor), c) if the rat shows epilation over the head where the radiation
was aimed, and d) if the rat shows any sign of abnormal behavior (including vocalization
upon touch). As of today, 45 days post-inoculation, three of the 15 rats are
alive and gaining weight. We plan to repeat the study, this time irradiating
the rats at day 14. Research was supported by grants from NIH, Georgetown University,
and from the brain tumor foundations of "Musella", "Lauren’s First
and Goal", and "Have a Chance".
Microbream Radiation Therapry: Applying The Method to Spinal Cord Research
in Rats. MEAGAN GREEN (La Salle University, Philadelphia, PA,
19141) F. AVRAHAM DILMANIAN (Brookhaven National Laboratory, Upton,
NY, 11973)
Presently there are no established
methods for stimulating the regeneration of the glial system in animals, yet
it is known that the system repairs itself after minor injuries. It has previously
been shown that the glial system recovers from irradiation with high dose planes
of thin synchrotron x rays. In this experiment we are interested in seeing if
irradiation treatment will not only cause recovery of the glial system but also
locomotive ability as well. Here we produced contusion injury in the rat spinal
cord and irradiated the site with arrays of parallel x-ray microplanar beams.
The New York University (NYU) impactor was used to produce contusions to the
T9 spinal region once the cord section was exposed by a laminectomy. The injury
temporarily paralyzed the rats from the T9 region down. The rats were treated
with antibiotics and were placed on heating pads to maintain a steady homeostasis
after surgery. They were then irradiated with arrays of thin, parallel x rays
(microbeams) at the National Synchrotron Light Source (NSLS). On the day before
the irradiations the rats were scored using the Basso et al scale to rank their
locomotor ability post surgery yet pre- irradiation. The range is 0-21, with
0 being the worst and fully paralyzed and 21 being the best and fully functional.
After the irradiations the rats were scored every four days to test how quickly
they were recovering. Within the first two weeks after irradiation the rats’ performance
appears to improve quickly, yet then it slows down drastically. We saw rapid
improvement within the first 14 days after irradiation. On the day of irradiation
two of the rats had a score of 3 and two of the rats had a score of 8. On day
14 after the initial irradiation, the two rats with the lower scores, achieved
a score of 9.5. The two rats with the higher scored achieved a score of 13. Because
of the lack of unirradiated controls in this primary study, the interpretation
of the results will have to wait until such controls are studied
and scored.
Molecular Cloning of Epitope-Tagged Mitochondria Fission and Fusion Genes. JOLY SHAH (Tallahassee Community College, Tallahassee,
FL, 32304) ANDREW GIFFORD (Brookhaven National Laboratory, Upton, NY,
11973)
There are two different types
of proteins in mammalian cells Drp-1 and fis-1 which are involved in mitochondria
fission. In contrast, there are three different types of proteins in mammalian
cells OPA-1, Mfn-1 and Mfn-2 which are essential for fusion of mitochondria.
But the central idea of this project is to make epitope tagged Drp1 or OPA1 DNA
constructs to study these proteins in cultured cells. Epitope are short stretch
of peptides that are antigenic. Proteins attached with epitope can be detected
by antibody that recognizes epitope. To make these constructs, we will use DNA
recombination and molecular cloning methods. Molecular cloning is the process
of generating genetically identical copies of DNA. We will first obtain cDNA
clone information of these genes will be purified by using Drp-1 and OPA-1 proteins,
in order to receive genetically identical copies of DNA fragments. It is also
necessary to use DNA recombination method to make epitope Flag tagged Drp-1 and
OPA-1 in pFLAG-CMV1 vector by growing some bacteria. In order to grow bacteria
Precision Scientific Thelco oven was use. To purified plasmid DNA the QIAprep
Spin miniprep kit and a centrifuge machine was use. Polymerase Chain Reaction
machine was also use in order to isolate fragment of DNA. Fisher Biotech machine
was use to find pure DNA and vector. By using this technique DNA was cut and
paste in vector to let it grow in clones. Moreover, molecular cloning book was
use for the more procedure. The project is still continuing, therefore the result
will indicate in future. In conclusion, cloning DNA fragment known as molecular
cloning. The purpose of growing DNA is to have large quantities of identical
copies of DNA for experiments.
National University Consortium Capability Catalog Development. MICHALENA
GROSSHANS (Ohio Northern University, Ada, OH, 45810) ROGER MAYES (Idaho
National Laboratory, Idaho Falls, ID, 83415)
The Idaho National Laboratory
(INL) established a National University Consortium (NUC) to enhance collaboration
with university faculty and further integrate university research and development
with INL programs. Five Academic Centers of Excellence (ACE), each with a different
focus that aligns with the INL mission, have also been established. The NUC
schools and the respective ACEs are as follows: Massachusetts Institute of
Technology (Advanced Reactor Fuels and Materials Technology), North Carolina
State University
(Simulation and Modeling), The Ohio State University
(Instrumentation & Control and Reactor Safety), Oregon State University (Thermal
Fluids and Reactor Safety) and the University of New Mexico (Non-Proliferation
Science and Technology). To facilitate collaboration between university and INL
researchers, the INL created a catalog of unique equipment, laboratories, projects,
software, computers, libraries etc., at each NUC facility, all connected to points
of contact. The catalog features data organized by Research Focus, Subcategory,
Keywords, Institution, Location, College, Department, Function/Capability, Description,
Resource Type, Collaborators/Sponsors/Partners, URL, Point of Contact including
phone number and e-mail, and Notes/Miscellaneous Information. The research foci
were identified by the ACEs and aligned with the laboratory’s strategic plan.
During this effort, information available on each university’s website was researched
thoroughly and gaps in the information were identified. Faculty and researchers
at each university have been asked to fill in the gaps. However, most requests
are pending response. The catalog is currently configured as an Excel spreadsheet
which will later provide INL researchers with an online, searchable database.
To maintain this catalog, continual updates will be needed to keep information
current. Future efforts will also capture the capabilities of the INL to make
the database useful for university researchers. Documentation to support these
efforts is available in the form of a user’s guide which provides insight as
to how the spreadsheet was generated and organized. This will allow someone else
to continue project development and potentially help end users improve the quality
of searches.
Optical Detection of Cocaine-induced Brain Function Changes in the Rat Brain
in vivo. MELISSA TULLY (State University of New York at Stony
Brook, Stony Brook, NY, 11790) CONGWU DU (Brookhaven National Laboratory,
Upton, NY, 11973)
Cocaine abuse increases the
risk of life-threatening neurological complications, has vasocontrictive properties
and also decreases the metabolism. However, the exact pathophysiological mechanisms
underlying cocaine’s neurotoxic effects remain incompletely understood. We have
developed a multi-wavelength spectroscope (MWS) to simultaneously measure the
changes in blood volume (CBV) and tissue oxygenation (StO2) from the brain surface.
We also have developed a Laser-Doppler Speckle Contrast Imaging (LDSCI) for detecting
the local cerebral blood flow (LCBF) changes in the living rat brain with a high
spatiotemporal resolution. In this study, we applied these new optical techniques
and combined them with Laser Doppler flowmetry (LDF) to systematically characterize
the effects of cocaine on cerebral blood flow (CBF), CBV, and StO2 as functions
of time in response to cocaine administration, as well to identify the spatial
feature of cocaine-induced LCBF changes in the somatosensory cortex of the living
brain. Three groups of rats were used in parallel to study the changes in CBF,
CBV and StO2, nd LCBF distribution in response to cocaine administration using
the optical systems of LDF, MWS and LDSCI, respectively. Each rat was anesthetized,
intubated, and mechanically ventilated, and the physiological parameters, including
the heart rate (ECG), respiration rate, mean arterial blood pressure (MABP) and
body temperature, were monitored. A craniotomy was then performed above the left
somatosensory cortex. An optical fiber-based probe, from MWS or LDF, or a CCD
camera of LDSCI was then mounted upon the exposed brain surface. The back-scattered
photons from the cortical surface were collected by the optical probe/camera
continuously before and after the cocaine administration. The changes of CBF,
CBV, StO2, and LCBF were determined as functions of time Our preliminary results
show that the cocaine induced a transient changes in CBF, CBV, and StO2. The
changes in CBF recovered to the baseline within 10-12 minutes whereas the changes
in CBV and StO2 in the brain were relatively longer lasting (> 20 minutes).
Interestingly, using the LDSCI system, the -chloralose anesthetized rats have
shown increased blood flow in large vessels and decreased flow in capillaries
or the microcirculation within the tissue bed in response to the cocaine administration.
These results explored the temporal and spatial features of the cerebrovascular
effects of cocaine on the brain, which might provide a better understanding of
etiology of cocaine related, stroke and transient ischemic attacks.
Patterning a PDMS Scaffold Towards Wound Healing. TEJAS
DOSHI (Wofford College, Spartanburg, SC, 29303) BARBARA BECKERMAN,
APRIL MCMILLAN, BOYD EVANS (Oak Ridge National Laboratory, Oak Ridge,
TN, 37831)
One of the most important
functions of skin is to act as a barrier against pathogens. This function is
compromised when a person suffers a partial-thickness or full-thickness wound.
The wound triggers a healing cascade that consists of three phases: inflammation,
proliferation, and tissue remodeling. The rate of wound healing determines
if scars form and how quickly skin can regain its purpose to serve as a barrier.
Depending on the severity of the wound, the healing process may last from a
few days to several months or longer. During the last thirty years, there has
been much interest and research in applying micro and nano-scaled technologies
to increase the rate of wound healing. These technologies include patterning
and functionalization of implantable materials. Patterning involves changing
the surface topography of a substrate. Functionalization involves attaching
different functional groups or signaling proteins to a substrate. The purpose
of this research is to determine a method to increase the rate of wound healing
using these micro and nano-scaled technologies. Extensive literature searches
were conducted to determine appropriate methods for patterning and functionalizing
a scaffold. Polydimethylsiloxane (PDMS) was chosen as the scaffold for the
experiments because it has been shown to be biocompatible with the human body.
Self-assembled polystyrene-block-polyacrylic acid (PS-b-PAA) diblock copolymer
micelles act as the template for patterning PDMS. The goal was to achieve regular,
evenly spaced patterns. However, the micelles formed bifurcated striations
which did not allow patterning of PDMS. This result is attributed to using
PS-b-PAA with a higher molecular mass than described in previous literature.
A close substitute was used because the vendor no longer sells the PS-b-PAA
described in the referenced experiment. Assuming that the micelles form regular,
evenly spaced patterns in future experiments, dermal fibroblasts will be seeded
onto molded scaffolds to determine if a patterned surface can best be used
to increase proliferation of skin cells so as to also increase the rate of
wound healing. Cell proliferation will be measured via a Coulter Counter device.
The substrate will also be functionalized with different signaling proteins
to induce a greater rate of wound healing. These experiments are the initial
stage of a long-term goal to create a therapeutic material that will increase
the rate of wound healing.
RS-GVG blocks methamphetamine-triggered reinstatement of the expression of
methamphetamine-induced conditioned place preference. JESSICA
PAI (New York University, New York, NY, 10012) STEPHEN DEWEY AND WYNNE
SCHIFFER (Brookhaven National Laboratory, Upton, NY, 11973)
This study investigated the
effects of RS-GVG on the reinstatement of the methamphetamine (METH)-induced
expression of conditioned place preference (CPP) in adolescent male Sprague-Dawley
rats. The expression of a CPP response correlating to dosage was first examined
by subjecting three groups of animals (n=8) to 30-minute pairings of METH (2.5,
5.0, and 10 mg/kg, respectively) with one chamber and 30-minute pairings of saline
in the other on alternate days over a 20-day period. Following completion of
the METH regimen, a CPP response was established. Subsequent daily testing identified
the rate of extinction for each group. On test days, animals received saline
and were permitted free access to both chambers for 20-minutes. The amount of
time spent in the METH-paired chamber was used to determine the conditioning
score. This test was performed until the expression of the CPP response was extinguished
(defined as 5 consecutive days without a significant CPP, p
Synthesis of Suberoyl Anilide Hydroxamic Acid for Positron Emission Tomography
Studies of its Inhibitory Effect in Histone. FRANK
OGERO (Medgar Evers College, Brooklyn, NY, 11225) KWESI AMOA (Brookhaven
National Laboratory, Upton, NY, 11973)
Suberoyl anilide hydroxamic
acid (SAHA) is a potent inhibitor of cell proliferation. Cell proliferation is
caused by the transcriptional role of histone deacetylase (HDAC) an enzyme that
influence tumorigenesis of cells. Our research involves synthesizing SAHA via
a two-step process. The initial step involved the reaction of suberic acid monomethyl
ester with aniline to afford methyl-8-anilino-8-oxooctanoate. This compound was
then treated with hydroxylamine to afford the desired compound. SAHA and (18Flouroacetamido)-1-hexanoic
acid (18FAHA), a radiotracer for HDAC, will be used to verify the Positron Emission
Tomography (PET) image with 18FAHA represents HDAC binding. More specifically,
if SAHA blocks the uptake of 18FAHA, this will serve as evidence that the image
represents HDAC in the brain. These studies will form the groundwork for future
PET studies in humans to understand the relationship between genes, brain chemistry
and behavior.
The Radiosynthesis of 6-(18Fluoroacetamido)-1-Hexanoicanilide for Positron
Emission Tomography Imaging of Histone Deacetylases in the Brain. SHANIKA
COLLINS (Medgar Evers College, Brooklyn, NY, 11207) JOANNA FOWLER (Brookhaven
National Laboratory, Upton, NY, 11973)
The substrate 6-(18Fluoroacetamido)-1-hexanoicanilide
(18F-FAHA) may be useful in measuring the level of histone deacetylase (HDAC)
expression and activity in cancer patients via Positron Emission Tomography (PET)
imaging and also in studying gene expression in the brain. Inhibition of HDACs
triggers growth arrest, differentiation and apoptosis in tumor cells. To evaluate
the HDAC expression in the brain and other organs in vivo, before and during
the use of inhibitors, FAHA was developed as a substrate for the HDAC to allow
for non-invasive whole body imaging of the HDAC. Our objective is to prepare
18F labeled FAHA to image HDAC in the brain in vivo using PET imaging. FAHA was
prepared by the reaction of 6-amino hexanoic acid with thionyl chloride in dichloroethane
followed by addition of aniline. The resulting product, compound 1, was then
treated with bromoacetyl bromide in the presence of triethylamine to afford compound
2 which was fluorinated using tetrabutylammonium fluoride to give our unlabeled
reference compound. The bromine substituted compound will serve as a precursor
to 18F-FAHAwhich will be used together with the HDAC inhibitor, suberoylanilide
hydroxamic acid for PET imaging of the HDAC expression and activity in the brains
of baboons. These studies will form the groundwork for future PET studies in
humans to understand the relationship between genes, brain chemistry and behavior.
The Role of Bootstrap Resampling to Improve Signal-to-Noise Ratio in PET Images. JOHN ZABROSKI (St. Joseph’s College, Patchogue, NY,
11772) JEAN LOGAN (Brookhaven National Laboratory, Upton, NY, 11973)
Positron Emission Tomography
(PET) imaging helps determine the effects of genetic variation, disease, behavior,
and drug administration on living systems at the cellular level. However, the
amount of information gained from these images is limited by noise due to poor
counting statistics related to the half-life of the radiotracer used and the
amount injected, which is limited by regulations placed on radiation exposure
to human subjects. For some radiotracers this noise also limits the ability to
create parametric images for each subject since an image is constructed by assigning
the biochemical parameter to each voxel. Group parametric images may overcome
these factors, eliminating error due to poor counting statistics by using discrete
time-frame averaging (DFA) before assigning the biochemical parameter to the
group image. However, the statistical significance that would otherwise be obtained
through individual parametric images cannot be determined with a group parametric
image, because there is only one image per group. To overcome this, bootstrap
re-sampling was used to create additional datasets-bootstrap samples. The initial
trial used data collected from the PET
radiotracer [11C]-Clorgyline (CLG), which binds to the enzyme monoamine oxidase
A (MAO-A). Simulated data was generated from the measured plasma input functions
using model parameters (K1, k2, k3) derived from a region of interest analysis of the thalamus
where k3 represents
binding of tracer to MAO-A and K1 and k2 represent transfer between plasma and tissue. By introducing
different levels of random noise to this data we can simulate time-activity curves
at the voxel level. Using DFA we then attempted to increase the signal-to-noise.
For each simulation, standard error was calculated. Also, each simulation was
compared to a bootstrapped standard error, with the number of bootstrap samples
necessary to obtain a stable estimate of the standard error systematically determined.
Our analysis shows that bootstrap resampling plays a useful role in determining
statistics associated with the DFA process. Bootstrap resampling was closest
to the true value of the voxel at the highest noise level (a=8.0). Now that we
have tested the validity of DFA in conjunction with bootstrap resampling, the
next step is to generate bootstrap parametric images, allowing an assessment
of the statistical significance of group differences.
Vigabatrin-Induced Weight Loss and Locomotor Sensitivity in Normal and Obese
Animals. STEFANIE AQUILINA (Cornell University, Ithaca, NY,
14853) DR. STEPHEN DEWEY (Brookhaven National Laboratory, Upton, NY,
11973)
Vigabatrin, an irreversible
inhibitor of GABA-transaminase, decreases dopamine in the central nervous system
(CNS). Recent studies suggest that much like drugs of abuse, food increases brain
dopamine in obese human volunteers. Combined with previous work using a host
of addictive substances, this study focused on an examination of food intake
and body weight in an animal model of obesity (Zucker rats). Dopamine is implicated
in behaviors associated with addictive drugs, including cocaine, nicotine, amphetamines,
alcohol, heroin, and a handful of illicit drug combinations. Unlike other prescription
drugs, short-term Vigabatrin use does not appear to have any significant and
potentially dangerous side effects. Two minor side effects were thought to include
weight fluctuations, as well as locomotor depression. Weight and food intake
in normal (adolescent Sprague Dawley rodents) and an animal model of obesity
(genetically altered adolescent and adult Zucker fatty rodents) were measured
during a period of two sub-chronic Vigabatrin administrations. In addition, locomotor
behavior boxes were used to analyze the activity in control animals and those
treated with Vigabatrin. Upon treatment with a high dose (150 mg/kg) of Vigabatrin
(active enantiomer) in normal rodents, significant weight loss was noted. Treatment
in obese rodents, however, resulted in a significantly slower rate of weight
gain. These weight loss trends appear dose-dependent (with more extreme weight
fluctuations occurring as a result of higher doses of Vigabatrin). However, locomotor
sensitivity was not significantly different between Vigabatrin treated and saline
treated animals. Thus, locomotor depression does not appear to be a side effect
of Vigabatrin. Furthermore, adult rodents treated daily with 300 mg/kg of Vigabatrin
(racemic) lost nearly 10% of their body weight in a period of only four days.
These studies suggest that Vigabatrin may be an effective treatment for obesity.
Additional studies at The University of Pennsylvania, University of California
Los Angeles, and Louisiana State University are currently ongoing in human subjects
using multiple doses of Vigabatrin and several treatment
protocols.
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