Cognitive deficits induced by 56Fe radiation exposure.

Exposing rats to particles of high energy and charge (e.g., 56Fe) disrupts neuronal systems and the behaviors mediated by them; these adverse behavioral and neuronal effects are similar to those seen in aged animals. Because cognition declines with age, and our previous study showed that radiation disrupted Morris water maze spatial learning and memory performance, the present study used an 8-arm radial maze (RAM) to further test the cognitive behavioral consequences of radiation exposure. Control rats or rats exposed to whole-body irradiation with 1.0 Gy of 1 GeV/n high-energy 56Fe particles (delivered at the alternating gradient synchrotron at Brookhaven National Laboratory) were tested nine months following exposure. Radiation adversely affected RAM performance, and the changes seen parallel those of aging. Irradiated animals entered baited arms during the first 4 choices significantly less than did controls, produced their first error sooner, and also tended to make more errors as measured by re-entries into non-baited arms. These results show that irradiation with high-energy particles produces age-like decrements in cognitive behavior that may impair the ability of astronauts to perform critical tasks during long-term space travel beyond the magnetosphere.     

The effects of proton exposure on neurochemistry and behavior.

Future space missions will involve long-term travel beyond the magnetic field of the Earth, where astronauts will be exposed to radiation hazards such as those that arise from galactic cosmic rays. Galactic cosmic rays are composed of protons, alpha particles, and particles of high energy and charge (HZE particles). Research by our group has shown that exposure to HZE particles, primarily 600 MeV/n and 1 GeV/n 56Fe, can produce significant alterations in brain neurochemistry and behavior. However, given that protons can make up a significant portion of the radiation spectrum, it is important to study their effects on neural functioning and on related performance. Therefore, these studies examined the effects of exposure to proton irradiation on neurochemical and behavioral endpoints, including dopaminergic functioning, amphetamine-induced conditioned taste aversion learning, and spatial learning and memory as measured by the Morris water maze. Male Sprague-Dawley rats received a dose of 0, 1.5, 3.0 or 4.0 Gy of 250 MeV protons at Loma Linda University and were tested in the different behavioral tests at various times following exposure. Results showed that there was no effect of proton irradiation at any dose on any of the endpoints measured. Therefore, there is a contrast between the insignificant effects of high dose proton exposure and the dramatic effectiveness of low dose (<0.1 Gy) exposures to 56Fe particles on both neurochemical and behavioral endpoints.     

CNS effects of heavy particle irradiation in space: behavioral implications.

Research from several sources indicates that young (3 mo) rats exposed to heavy particle irradiation (56Fe irradiation) produces changes in motor behavior as well as alterations in neuronal transmission similar to those seen in aged (22-24 mo) rats. These changes are specific to neuronal systems that are affected by aging. Since 56Fe particles make up approximately 1-2% of cosmic rays, these findings suggest that the neuronal effects of heavy particle irradiation on long-term space flights may be significant, and may even supercede subsequent mutagenic effects in their mission capabilities. It is suggested that among other methods, it may be possible to utilize nutritional modification procedures to offset the putative deleterious effects of these particles in space.     

Effects of heavy particle irradiation and diet on object recognition memory in rats

On long-duration missions to other planets astronauts will be exposed to types and doses of radiation that are not experienced in low earth orbit. Previous research using a ground-based model for exposure to cosmic rays has shown that exposure to heavy particles, such as ⁵⁶Fe, disrupts spatial learning and memory measured using the Morris water maze. Maintaining rats on diets containing antioxidant phytochemicals for 2 weeks prior to irradiation ameliorated this deficit. The present experiments were designed to determine: (1) the generality of the particle-induced disruption of memory by examining the effects of exposure to ⁵⁶Fe particles on object recognition memory; and (2) whether maintaining rats on these antioxidant diets for 2 weeks prior to irradiation would also ameliorate any potential deficit. The results showed that exposure to low doses of ⁵⁶Fe particles does disrupt recognition memory and that maintaining rats on antioxidant diets containing blueberry and strawberry extract for only 2 weeks was effective in ameliorating the disruptive effects of irradiation. The results are discussed in terms of the mechanisms by which exposure to these particles may produce effects on neurocognitive performance.     

Long-term changes in amphetamine-induced reinforcement and aversion in rats following exposure to 56Fe particle.

Exposing rats to heavy particles produces alterations in the functioning of dopaminergic neurons and in the behaviors that depend upon the integrity of the dopaminergic system. Two of these dopamine-dependent behaviors include amphetamine-induced reinforcement, measure using the conditioned place preference procedure, and amphetamine-induced reinforcement, measured using the conditioned place preference procedure, and amphetamine-induced aversion, measured using the conditioned taste aversion. Previous research has shown that exposing rats to 1.0 Gy of 1GeV/n 56Fe particles produced a disruption of an amphetamine-induced taste aversion 3 days following exposure, but produced an apparent enhancement of the aversion 112 days following exposure. The present experiments were designed to provide a further evaluation of these results by examining taste aversion learning 154 days following exposure to 1.0 Gy 56Fe particles and to establish the convergent validity of the taste aversion results by looking at the effects of exposure on the establishment of an amphetamine-induced conditioned place preference 3, 7, and 16 weeks following irradiation. The taste aversion results failed to confirm the apparent enhancement of the amphetamine-induced CTA observed in the prior experiment. However, exposure to 56Fe particles prevented the acquisition of amphetamine-induced place preference at all three-time intervals. The results are interpreted as indicating that exposure to heavy particles can produce long-term changes in behavioral functioning.     

CNS-induced deficits of heavy particle irradiation in space: the aging connection.

Our research over the last several years has suggested that young (3 mo) rats exposed to whole-body 56Fe irradiation show neuronal signal transduction alterations and accompanying motor behavioral changes that are similar to those seen in aged (22-24 mo) rats. Since it has been postulated that 1-2% of the composition of cosmic rays contain 56Fe particles of heavy particle irradiation, there may be significant CNS effects on astronauts on long-term space flights which could produce behavioral changes that could be expressed during the mission or at some time after the return. These, when combined with other effects such as weightlessness and exposure to proton irradiations may even supercede mutagenic effects. It is suggested that by determining mechanistic relationships that might exist between aging and irradiation it may be possible to determine the common factor(s) involved in both perturbations and develop procedures to offset their deleterious effects. For example, one method that has been effective is nutritional modification.     

Behavioral endpoints for radiation injury.

The relative behavioral effectiveness of heavy particles was evaluated. Using the taste aversion paradigm in rats, the behavioral toxicity of most types of radiation (including 20Ne and 40Ar) was similar to that of 60Co photons. Only 56Fe and 93Nb particles and fission neutrons were significantly more effective. Using emesis in ferrets as the behavioral endpoint, 56Fe particles and neutrons were again the most effective; however, 60Co photons were significantly more effective than 18 MeV electrons. These results suggest that LET does not completely predict behavioral effectiveness. Additionally, exposing rats to 10 cGy of 56Fe particles attenuated amphetamine-induced taste aversion learning. This behavior is one of a broad class of behaviors which depends on the integrity of the dopaminergic system and suggests the possibility of alterations in these behaviors following exposure to heavy particles in a space radiation environment.     

Effects of exposure to heavy particles on a behavior mediated by the dopaminergic system.

The effects of exposure to heavy particles on behaviors mediated by the central nervous system (CNS) are qualitatively different than the effects produced by exposure to other types of radiation. One behavior mediated by the CNS is the amphetamine-induced taste aversion, which is produced by pairing a novel tasting solution with injection of amphetamine. When the conditioning day is three days following irradiation, exposing rats to low doses of 56Fe particles (600 MeV/n or 1 GeV/n) eliminates the taste aversion produced by injection of amphetamine, which is dependent upon the integrity of the central dopaminergic system, but has no effect on the aversion produced by injection of lithium chloride which is mediated by the gastrointestinal system. In contrast to the effects obtained using heavy particles, exposing rats to 60CO gamma rays or to fission spectrum neutrons has no selective effect upon the acquisition of either amphetamine- or lithium chloride-induced taste aversions. When the conditioning day occurs four months following exposure to 1 GeV/n 56Fe particles, there is an enhancement of the amphetamine-induced taste aversion. The implications of these findings for approaches to risk assessment are considered.     

Morphometric studies of heavy ion damage in the brains of rodents.

As an approach to determining the relative biological effectiveness (RBE) of each of five different heavy ions for the mammalian brain, histological preparations of brains from mice exposed to various HZE particles at different doses and primary LETinfinity values were examined by means of semi-automated image analysis for volume changes in specific regions of the olfactory bulb. The mice were irradiated at 100 days of age and euthanatized about 500 days (16 months) later. Exposures were: 60Co gamma photons (LETinfinity = 1-2 keV/micrometer), 4He (LETinfinity = 6 keV/micrometer), 12C (LETinfinity = 80 keV/micrometer), 20Ne (LETinfinity = 150 keV/micrometer), 56Fe (LETinfinity = 180 keV/micrometer), and 40Ar (LETinfinity = 650 keV/micrometer). Animals receiving particle radiation were exposed in an extended Bragg peak region except for iron where the plateau region was used. The zones measured in the olfactory bulb were 1) the external plexiform layer (zone) and 2) an internal region consisting of the granule cells, internal plexiform layer, and layer of mitral cells. These studies indicated that volume changes did indeed occur, not only in absolute terms but also when expressed as the ratio of the structures to each other and to the bulb as a whole. Although this study is exploratory in character, the data obtained may nevertheless contribute to a determination of risk factors due to late effects from HZE articles.     

The influence of dose, dose-rate and particle fragmentation on cataract induction by energetic iron ions.

Because activities in space necessarily involve chronic exposure to a heterogeneous charged particle radiation field it is important to assess the influence of dose-rate and the possible modulating role of heavy particle fragmentation on biological systems. Using the well-studied cataract model, mice were exposed to plateau 600 MeV/amu 56Fe ions either as acute or fractionated exposures at total doses of 5 - 504 cGy. Additional groups of mice received 20, 360 and 504 cGy behind 50 mm of polyethylene, which simulates body shielding. The reference radiation consisted of 60Co gamma radiation. The animals were examined by slit lamp biomicroscopy over their three year life spans. In accordance with our previous observations with heavy particles, the cataractogenic potential of the 600 MeV/amu 56Fe ions was greater than for low-LET radiation and increased with decreasing dose relative to gamma-rays. Fractionation of a given dose of 56Fe ions did not reduce the cataractogenicity of the radiation compared to the acute regimen. Fragmentation of the beam in the polyethylene did not alter the cataractotoxicity of the ions, either when administered singly or in fractions.     

Effects of exposure to different types of radiation on behaviors mediated by peripheral or central systems.

The effects of exposure to ionizing radiation on behavior may result from effects on peripheral or on central systems. For behavioral endpoints that are mediated by peripheral systems (e.g., radiation-induced conditioned taste aversion or vomiting), the behavioral effects of exposure to heavy particles (56Fe, 600 MeV/n) are qualitatively similar to the effects of exposure to gamma radiation (60Co) and to fission spectrum neutrons. For these endpoints, the only differences between the different types of radiation are in terms of relative behavioral effectiveness. For behavioral endpoints that are mediated by central systems (e.g., amphetamine-induced taste aversion learning), the effects of exposure to 56Fe particles are not seen following exposure to lower LET gamma rays or fission spectrum neutrons. These results indicate that the effects of exposure to heavy particles on behavioral endpoints cannot necessarily be extrapolated from studies using gamma rays, but require the use of heavy particles.     

Head and neck tumors after energetic proton irradiation in rats.

This is a two-year progress report on a life span dose-response study of brain tumor risk at moderate to high doses of energetic protons. It was initiated because a joint NASA/USAF life span study of rhesus monkeys that were irradiated with 55-MeV protons (average surface dose, 3.5 Gy) indicated that the incidence of brain tumors per unit surface absorbed dose was over 19 times that of the human tinea capitis patients whose heads were exposed to 100 kv x-rays. Examination of those rats that died in the two-year interval after irradiation of the head revealed a linear dose-response for total head and neck tumor incidence in the dose range of 0-8.5 Gy. The exposed rats had a greater incidence of pituitary chromophobe adenomas, epithelial and mesothelial cell tumors than the unexposed controls but the excessive occurrence of malignant gliomas that was observed in the monkeys was absent in the rats. The estimated dose required to double the number of all types of head and neck tumors was 5.2 Gy. The highest dose, 18 Gy, resulted in high mortality due to obstructive squamous metaplasia at less than 50 weeks, prompting a new study of the relative biological effectiveness of high energy protons in producing this lesion.     

Early and delayed reproductive death in human cells exposed to high energy iron ion beams.

The aim of this research was to determine the biological effectiveness for early and delayed effects of high energy, high linear energy transfer (LET) charged particles. Survival and delayed reproductive death were measured in AG1522 human fibroblast cells exposed to Fe-ion beams of energies between 0.2 and 1 GeV/n, 0.97 GeV/n Ti-ion and 0.49 GeV/n Si-ion beams. The cells were irradiated at the HIMAC accelerator in Chiba, Japan (0.2 and 0.5 GeV/n Fe and 0.49 GeV/n Si) and at the NASA Space Radiation Laboratory in Brookhaven, USA (1 GeV/n Fe and 0.97 GeV/n Ti ions). The dose-effect curves were measured in the dose range between 0.25 and 2 Gy. For comparison cells were exposed to 60Co gamma rays. Analysis of the dose-effect curves show that all the heavy ion beams induce inactivation and delayed reproductive death more effectively than 60Co gamma rays. The only exception is the 0.2 GeV/n Fe-ion beam at low doses. The progeny of the irradiated cells show delayed damage in the form of reproductive death with all the heavy ion beams with the 1 GeV/n Fe-ion beam being the most effective. The relative biological effectiveness at low doses of the iron beams is highest for LET values between 140 and 200 keV/micrometers with values of 1.6 and 3 for early and delayed reproductive death, respectively. Analysis of the fluence-effect curves shows that the cross-sections for early and delayed inactivation increase with increasing LET up to 442 keV/micrometers.     

Potential vascular damage from radiation in the space environment.

Cultured endothelial cells of blood vessels have a Do of 2 Gy for X-rays. A dose of 0.5 Gy of X-rays has an acute effect on vessel diameter. The vessels may show other acute effects such as change in permeability including a change in the blood brain barrier. Changes occurring from late effects of chronic exposure in vascular architecture include telangiectasia and decrease in vascular density. Changes in the perivascular connective tissue particularly collagen may play a role in these changes. After charged particle exposure of 15 and 30 Gy, radiation changes in the blood brain barrier and vascular changes are noted in the nervous system. These long term changes are recorded by PET, MRI, and CT imaging. Chronic exposure to alpha particles causes vascular damage in compact bone resulting in bone infarcts. Using tandem scanning confocal microscopy in-situ imaging of the capillaries and collagen of the papillary dermis provides a non-invasive method of serial recording of changes in irradiated microvasculature.     

Effects of 40Ar and 56Fe ions on retinal photoreceptor cells of the rabbit: implications for manned missions to Mars.

Losses of photoreceptor cells (rods) from the retinas of New Zealand white (NZW) rabbits were detectable within 2 years after localized acute irradiation of optic and proximal tissues with > or = 7 Gy of 530 MeV u-1 40Ar ions or > or = 2 Gy of 465 MeV u-1 56Fe ions in the Bragg plateau region of energy deposition. Those limits were determined only from an analysis of variance of dose groups because the shapes of the dose response curves at early post-irradiation times are not known, a concern being addressed by experiments in progress. Losses of photoreceptor cells for the period 0.5-2.5 years post-irradiation, determined by provisional linear regression analysis, were approximately 1.7% Gy-1 and 2.5% Gy-1 for 40Ar and 56Fe ions, respectively.     

Operant responding following exposure to HZE particles and its relationship to particle energy and linear energy transfer

On exploratory class missions astronauts will be exposed to a variety of heavy particles (HZE particles) which differ in terms of particle energy and particle linear energy transfer. The present experiments were designed to evaluate how these physical characteristics of different particles affect cognitive performance, specifically operant responding. Following exposure to ²⁸Si, ⁴⁸Ti, ¹²C and ¹⁶O particles at the NASA Space Radiation Laboratory rats were tested for their ability to respond appropriately to changes in reinforcement schedules using an operant task. The results showed that the effectiveness of different particles in disrupting cognitive performance, defined as the lowest dose that produced a performance decrement, varied as a function of the energy of the specific particle: for comparisons between different energies of the same particle (e.g., ⁵⁶Fe) the effectiveness of the particle was directly proportional to particle linear energy transfer, whereas for comparisons between different particles (e.g., ⁵⁶Fe and ¹⁶O) effectiveness was inversely proportional to particle linear energy transfer. The results are discussed in terms of the mechanisms that influence the effectiveness of different particles and energies and in terms of their implications for analyzing the possible risks to astronauts of decrements in cognitive performance following exposure to HZE particles on long-duration exploratory class missions.     

Microlesions: theory and reality.

Efforts to assess radiation risk in space have been complicated by the considerable unknowns regarding the biological effects of the heavy ion component (HZE particles) of the cosmic rays. The attention has focused primarily on the assignation of a quality factor (Q) which would take into account the greater effectiveness of heavy ions vis-a-vis other forms of ionizing radiation. If however, as the so-called "Microlesion Theory" allows, the passage of HZE particles through living tissue produces unique biological damage, the traditional use of Q becomes meaningless. Therefore, it is critical to determine if microlesions, in fact, do exist. While the concept does not necessarily require detectable morphological damage, "tunnel-lesions" or holes in ocular tissues have been cited as evidence of microlesions. These data, however, are open to reinterpretation. On-going light, scanning and transmission electron microscopic studies of the corneas, lenses and retinas of rat eyes exposed to 450 MeV/amu 56Fe ions thus far have not revealed tunnel-lesion damage. The morphological effects of the heavy ions have been found to be qualitatively similar to the changes following other kinds of ionizing radiation.     

Induction of genomic instability after an acute whole-body exposure of mice to Fe ions

The purpose of this study was to evaluate dose–response relationships for the in vivo induction of micronuclei (MN) as a measure of both initial radiation damage and the induction of genomic instability. These measurements were made in mouse blood erythrocytes as a function of radiation dose, radiation quality, time after irradiation, and the genetic background of exposed individuals. Blood samples were collected from two strains of mouse (CBA/CaJ and C57BL/6J) at different times up to 3 months following a whole-body exposure to various doses of 1 GeV/amu ⁵⁶Fe ions (0, 0.1, 0.5 and 1.0 Gy, at the dose rate of a 1 Gy/min) or ¹³⁷Cs gamma rays (0, 0.5, 1.0 and 3.0 Gy, at the dose rate of 0.72 Gy/min). Blood-smear slides were stained with acridine orange (AO). The frequencies of MN were measured in mature normochromatic-erythrocytes (MN-NCEs) and in immature polychromatic-erythrocytes (MN-PCEs). Effects of both types of radiation on erythropoiesis were also evaluated. As a measure of cell progression delay, a dose-dependent decrease in numbers of PCEs was observed at day 2 post-exposure in both strains, regardless of radiation quality. Subsequently, the levels of PCEs increased in all exposed mice, reaching control levels (or higher) by day 7 post-exposure. Further, at day 2 after the exposure, there was no increase in the frequency of MN-PCEs in CBA/CaJ mice exposed to ⁵⁶Fe ions while the frequency of MN-PCEs elevated as a function of dose in the C57BL/6J mice. At day 4, there was no dose related increase in MN-NCEs in either strain of mouse exposed to ¹³⁷Cs gamma rays. Additionally, at the early sacrifice times (days 2 and 4), ⁵⁶Fe ions were slightly more effective (per unit dose) in inducing MN-NCEs than ¹³⁷Cs gamma rays in CBA/CaJ mice. However, there was no increase in the frequency of MN-NCEs at late times after an acute exposure to either type of radiation. In contrast, both types of radiation induced increased MN-PCEs frequencies in irradiated CBA/CaJ mice, but not C57BL/6J mice, at late times post-exposure. This finding indicates the potential induction of genomic instability in hematopoietic cells of CBA/CaJ mice by both types of radiation. The finding also demonstrates the influence of genetic background on radiation-induced genomic instability in vivo.     

Oncogenic transformation of mammalian cells by ultrasoft X-rays and alpha particles.

For a better understanding of oncogenic cell transformation by ionizing radiation, we conducted experiments with ultrasoft X rays and low energy alpha particles. Confluent C3H10T1/2 cells were irradiated by Al-K (1.5 keV) X rays or alpha particles from plutonium through a thin mylar sheet, on which the cells attached and grew. Our results indicated that Al-K X rays were more effective in causing cell inactivation and oncogenic transformation than 60Co gamma rays but less effective than 1.0 and 3.7 MeV alpha particles. There was no significant difference between 1.0 and 3.7 MeV alpha particles in transforming cells although the latter were slightly more effective than the former in producing lethal effect. These results indicated that track structure is important in causing biological effects by ionizing radiation.     

Histologic effects of high energy electron and proton irradiation of rat brain detected with a silver-degeneration stain.

Application of the degeneration sensitive, cupric-silver staining method to brain sections of male Sprague-Dawley rats irradiated 4 days before sacrifice with 155 Mev protons, 2-8 Gy at 1 Gy/min (N=6) or 22-l0lGy at 20 Gy/min (N=16) or with 18.6 Mev electrons, 32-67 Gy at 20 Gy/min (N=20), doses which elicit behavioral changes (accelerod or conditioned taste aversion), resulted in a display of degeneration of astrocyte-like cell profiles which were not uniformly distributed. Plots of 'degeneration scores' (counts of profiles in 29 areas) vs. dose for the proton and electron irradiations displayed a linear dose response for protons in the range of 2-8 Gy. In the 20-100 Gy range, for both electrons and protons the points were distributed in a broad band suggesting a saturation curve. The dose range in which these astrocyte-like profiles becomes maximal corresponds well with the dose range for the X-ray eradication of a subtype of astrocytes, 'beta astrocytes'.