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Project Title:  Functional decline in mice with Alzheimer's-type neurodegeneration is accelerated by charge-particle radiation Reduce
Fiscal Year: FY 2015 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 02/01/2011  
End Date: 01/31/2015  
Task Last Updated: 04/07/2015 
Download report in PDF pdf
Principal Investigator/Affiliation:   Vlkolinsky, Roman  Ph.D. / Loma Linda University 
Address:  11175 Campus St 
Chan Shun Pavilion, A-1010 
Loma Linda , CA 92350-1700 
Email: rvlkolinsky@llu.edu 
Phone: 909-558-7403  
Congressional District: 41 
Web:  
Organization Type: UNIVERSITY 
Organization Name: Loma Linda University 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Nelson, Gregory  Ph.D. Loma Linda University 
Hartman, Richard  Ph.D. Loma Linda University 
Key Personnel Changes / Previous PI: Jerome Badaut, PhD terminated participation in our project as of July, 2013. Richard E Hartman, PhD ; Gregory Nelson, PhD ; Attila Szucs, PhD - subcontractor
Project Information: Grant/Contract No. NNX11AE41G 
Responsible Center: NASA JSC 
Grant Monitor: Simonsen, Lisa  
Center Contact:  
lisa.c.simonsen@nasa.gov 
Solicitation / Funding Source: 2010 Space Radiobiology NNJ10ZSA001N 
Grant/Contract No.: NNX11AE41G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) CNS:Risk of Acute (In-flight) and Late Central Nervous System Effects from Radiation Exposure (IRP Rev G)
Human Research Program Gaps: (1) CNS02:Does space radiation exposure elicit key events in adverse outcome pathways associated with neurological diseases? What are the key events or hallmarks, their time sequence and their associated biomarkers? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: End date is now 1/31/2015 per NSSC information (Ed., 11/5/13)

Task Description: Exposure of an astronaut’s central nervous system (CNS) to solar particle events (SPE) and galactic cosmic rays (GCR) may accelerate neurodegenerative changes and impact neuronal network activity, leading to cognitive deficits. There are similarities between radiation-induced CNS effects and pathological processes found in the Alzheimer’s disease (AD). Common functional and structural findings include profound deficits in neuronal communication (synaptic transmission), cognitive impairments, neuro-inflammatory changes, and reduced neurogenesis. These similarities lead us to hypothesize that subjects with a genetic propensity to develop AD-pathology may be excessively vulnerable to ionizing radiation. We previously showed in transgenic (TG) APP23 mice, a murine model of AD, that irradiation with 600 MeV/n iron particles accelerated the onset of electrophysiological changes in the hippocampus, a brain structure crucially involved in the formation of short-term memory. In this project we use young adult APP/PSEN1E9 (APP/PSEN1) double transgenic (TG) mice and expose them to low doses of 150 MeV proton (irradiations performed at LLU proton treatment facility), 250 MeV/n silicon and 600 MeV/n iron particle radiation to compare and quantify their detrimental effects on hippocampal functions and the onset of AD-like pathology. The APP/PSEN1 TG mice typically exhibit early-onset of age-related behavioral abnormalities and deficits in synaptic transmission. We hypothesized that exposure to even low radiation doses will accelerate the onset of age-related neurodegenerative processes, while in wild-type (WT) animals such damage may stay undetectable. Comparison of proton, silicon, and iron radiation on selected neurophysiological end points in APP/PSEN1 TG mice will provide valuable information whether exposure to space radiation may exacerbate neurodegenerative processes. The functional end points (e.g., electrophysiological and behavioral changes) will be directly correlated with the expression of immuno-histochemical markers of neurodegeneration, including amyloid plaque load, synaptic proteins, and expression of neuro-inflammatory cytokines. If such correlation is found, it may indicate causative relationship between decrements in hippocampal functions and structural changes, which will help to elucidate the pathological mechanisms of radiation-induced neuronal injury and estimate the risks of low dose HZE radiation exposure to the CNS.

Research Impact/Earth Benefits: The central nervous system (CNS) has been typically described as radiation-resistant tissue. However, there is now sufficient body of evidence, mostly from experiments in rodents, showing that low doses of charged particle radiation (< 1 Gy) may affect some basic neuronal processes, such as synaptic excitability, neuronal firing and propensity for epileptiform activity. In addition, it has been posited by us and other investigators that in subjects prone to develop neurodegenerative diseases such as Alzheimer Disease (AD), an exposure to charged-particle radiation may accelerate the onset and/or alter the progression of AD, increase amyloid plaque load, and promote neuro-inflammatory changes within their brain. While such effects have been observed with high-LET particles (e.g., 1GEv/n Iron nuclei), this hypothesis has not been fully tested with other low- and high-LET particles and protons that represent major components in space radiation spectra. Studying the impact of protons and high-LET radiation on neurodegenerative processes in mammalian CNS is a critical step for critical assessment of the space radiation CNS-risks for astronauts, and e.g. for further development of modern cranial radiotherapies using charged particle radiation. The time-dependent changes in the CNS in patients undergoing cranial irradiations have been well documented, and they range from mild memory deficits to severe delayed demyelination and neurodegeneration. Whether low doses of charged particle radiation may accelerate the onset or affect the severity of AD-related pathology is not known. In the current project we used a murine double transgenic (TG) model of AD (the APP/PSEN1 TG mice commercially available from Jackson Laboratories) that we exposed to low- and high-LET charged-particle radiation and tested whether radiation affects the time course and severity of neurodegenerative processes in these TG mice. The functional changes were also compared to limited cohort of wild type (WT) mice to rule out qualitative differences in response to irradiation. The combination of behavioral, electrophysiological, and histological techniques helped us to gather unique data on radiation effects from 150 MeV proton-, 250 MeV/n silicon-, and 600 MeV/n iron-irradiated APP/PSEN1 TG mice. We identified multitude of functional changes in WT and TG brain and cortex of these mice, such as changes in synaptic excitability in the hippocampal CA1 neurons, and altered short term synaptic plasticity. Interestingly, in WT mice we observed radiation-induced suppression of epileptiform activity, a finding that may be clinically relevant in reducing epileptic seizures. In TG mice irradiated with protons and HZE, we observed altered presynaptic proteins involved in neurotransmitter release and worsened neurodegenerative changes. The acquired data will improve our understanding of pathophysiological processes in irradiated and AD-affected CNS tissue and will help to assess CNS-radiation risks for future manned, deep-space missions, such as the mission to Mars.

Task Progress & Bibliography Information FY2015 
Task Progress: Aim 1 & Aim 3 Activities. In accord with our statement of work, we completed all irradiations, behavioral testing, and in vitro electrophysiological experiments with protons and HZE (silicon 250 MeV/n & iron 600 MeV/n nuclei) particles. Proton irradiated APP/PSEN1 transgenic (TG) and wild-type (WT) mice were behaviorally tested pre- and 3 and 6 months post-irradiation followed by electrophysiological testing either at 6 or 9 months post-irradiation, as planned. HZE-irradiated mice (TG only) were behaviorally tested 3 and 6 months post-irradiation, followed by electrophysiological testing at 6-7 months post-irradiation. In total, from 2010-2013 we irradiated 78 TG and 16 WT mice with protons at Loma Linda University, Proton Treatment Facility. Electrophysiological testing was successfully performed in 82 proton-irradiated mice. We irradiated 120 TG with HZE particles at NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratories. Seven TG mice died spontaneously, thus electrophysiological testing was successfully performed on 113 animals. We used conventional extracellular recordings to monitor evoked synaptic responses and spontaneous epileptiform activity. The analyses of behavioral and electrophysiological data from all irradiated animals, including statistical evaluations, have been completed. We report that at radiation doses ranging from 0.1-1 Gy, whole–body exposures to protons, silicon or iron nuclei did not affect survival of TG mice.

Behavioral analyses performed in cohorts of mice irradiated with protons (specific Aim 1) using water maze (WM), Barnes maze (BM), and zero maze (ZM) confirmed previously described, Alzheimer disease (AD) genotype-related deficits in spatial memory that were likely associated with hippocampal dysfunction. Genotype-related behavioral decrements were evident already at 3 months post-irradiation. For example, in control (non-irradiated) APP/PSEN1 TG mice we observed an increased swim distance to the target area in WM, an increased average distance moved in BM, and a reduction of time spent in the dark side of the ZM, when compared to the control (0 Gy) WT mice. Such findings indicate AD-related decrements in spatial memory (WM and BM data), but paradoxically a reduced anxiety-like behavior (ZM data) in TG mice.

We observed relatively subtle, proton- and HZE-radiation-induced behavioral effects. Significant radiation-induced effects were only observed in WT mice after exposure to 0.5 Gy protons detected in WM at 6 months post-irradiation. In these mice only (n=16), we observed significantly increased swim distance during reversal learning phase of the WM test, indicating reduced cognitive flexibility. While proton radiation (tested at 0.5 Gy only) affected the performance of WT mice, exposures to protons and HZE-radiation (tested at 0.1-1 Gy) did not affect the performance of APP/PSEN1 TG mice. Such result may indicate that low doses of proton and HZE radiation may not necessarily worsen the functional outcomes in mice prone to AD-like pathology, or alternatively such pathology may trump any observable behavioral radiation-induced effects. Interestingly, the TG mice irradiated with 250 MeV/n silicon particles exhibited reduced performance in WM at 3 months only; the decrement was reaching statistical significance at 0.1 Gy only and appeared to be transient as it could not be detected at 6 months post-irradiation. No significant differences were observed for either HZE species in BM and ZM. In TG mice irradiated with 600 MeV/n iron particles we surprisingly observed a trend for improved performance in WM (reduced cumulative distance to the target platform), at 6 months post-irradiation at the dose of 1 Gy.

Electrophysiological data indicate that proton radiation at doses from 0.1 to 1 Gy may impact synaptic excitability and short-term synaptic plasticity mediated by presynaptic glutamate release, but it likely does not affect long-term potentiation (LTP; reported previously to be altered in HZE-irradiated mice), the widely used cellular correlate of memory formation in the hippocampus. We observed that proton radiation-induced changes in synaptic excitability are qualitatively different in TG when compared to changes observed in WT mice. In accord with our behavioral findings, the WT mice exhibit different sensitivity to radiation and, for example at 0.5 Gy, we observed increased postsynaptic excitability in CA1 neurons, whereas the TG mice exhibited opposite responses at the same radiation dose. The stimulation paradigms testing the effect of proton radiation on presynaptic glutamate release (paired-pulse facilitation; PPF) revealed that the TG mice (but not the WT mice) at 9 exhibited reduced PPF indicating increased glutamate release. In addition, only in the WT mice irradiated with 0.5 Gy we observed reduced incidence of epileptiform activity (tested in the CA3-CA1 hippocampal network) promoted by activation of NMDA receptors. Interestingly, in TG mice, a radiation exposure to protons or HZE particles had no effect on these spontaneous oscillations.

Aim 2 Activities. We completed immunohistological evaluations of ß-amyloid deposits in the brain samples (the cortex and the hippocampus) of APP/PSEN1 TG mice irradiated with protons and HZE particles using 6E10 monoclonal antibody (total amyloid). We confirmed an amyloid depositions in all brains of APP/PSEN1 TG mice at 6 and 9 months post-irradiation time point. At 9 months, we detected proton radiation-induced significant increases of total amyloid deposition at 1Gy in the dorsal cortex, but not the hippocampus. However, we did not identify similar increases in plaque deposition in TG mice irradiated with silicon or iron nuclei.

Neuro-inflammation and neurodegenerative changes in TG (and WT) brains (cortex only) exposed to radiation have been assessed by analyses of five cytokines/chemokines (IL-1b, IL-6, TNFa, MCP-1, and IL-10) in homogenates of TG and WT mouse cortices. These signaling molecules have been previously reported to be elevated in irradiated brains and/or have been shown to affect synaptic plasticity in the hippocampus, thus their elevation may be associated with functional decrements observed in these animals. In a cohort of proton-irradiated mice we observed differences in the expression of chemokine IL-10 (CXCL10) between TG and WT mice at 9 months, but the effect was not dependent on the radiation exposure. The other chemokines were not affected by either genotype or radiation, indicating that at 9 months radiation effects on the CNS are not associated with elevated levels of pro-inflammatory cytokines. This also indicated that the electrophysiological and behavioral decrements reported above are not due to elevated levels of cytokines within the CNS, as previously suggested by us and other investigators.

We finished analyses of synaptic marker synaptophysin in cortices of WT and TG mice irradiated with protons and HZE radiation by Western blotting (WB). In WT mice at 9 months post-irradiation with 0.5 Gy protons, we observed significantly increased synaptophysin expression when compared to control WT mice. Interestingly, in all TG mice groups (control 0.5 and 1 Gy only) the synaptophysin levels were comparable to those found in irradiated WT mice reflecting presumably an AD-related pathology (a genotype effect). However, the proton-irradiation in TG mice had no further enhancing effects on this presynaptic marker. Thus, we suggest that proton radiation does not affect an elevated expression of synaptophysin in subjects prone to AD-pathology. WB analyses of synaptophysin expression performed in the Si- and Fe-irradiated APP/PSEN1 TG mice at 6-7 mo post-irradiation indicated significant radiation-induced changes. While the Si-irradiated mice exhibited trends of increased expression at 0.1 and 0.5 Gy and reduction to control levels at 1 Gy, the Fe-irradiated mice exhibited significant reduction at 1 Gy.

Several manuscripts are in preparation for submission to peer-reviewed journals.

Bibliography Type: Description: (Last Updated: 04/24/2019) 

Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Rudobeck E, Szucs A, Vlkolinsky R. "Effects of proton radiation on evoked and spontaneous neuronal activity in the hippocampus of APP/PSEN1 transgenic mice." HITSRS2013--Heavy Ion in Therapy and Space Radiation Symposium 2013, Chiba, Japan, May 15-18, 2013.

J Radiat Res. 2014 Mar;55 (Suppl 1):i102-i103. http://dx.doi.org/10.1093/jrr/rrt174 ; Extended abstract. , Mar-2014

Abstracts for Journals and Proceedings Bellone JA, Hartman RE, Vlkolinsky R. "The effects of low doses of proton, iron or silicon radiation on spatial learning in a mouse model of Alzheimer's disease." HITSRS2013--Heavy Ion in Therapy and Space Radiation Symposium 2013, Chiba, Japan, May 15-18, 2013.

J Radiat Res. 2014 March 55(Suppl 1):i95–i96. http://dx.doi.org/10.1093/jrr/rrt154 ; Extended abstract. , Mar-2014

Articles in Peer-reviewed Journals Bellone JA, Rudobeck E, Hartman RE, Szücs A, Vlkolinský R. "A single low dose of proton radiation induces long-term behavioral and electrophysiological changes in mice." Radiat Res. 2015 Aug;184(2):193-202. https://doi.org/10.1667/RR13903.1 ; PubMed PMID: 26207690 , Aug-2015
Articles in Peer-reviewed Journals Rudobeck E, Bellone JA, Szücs A, Bonnick K, Mehrotra-Carter S, Badaut J, Nelson GA, Hartman RE, Vlkolinsky R. "Low-dose proton radiation effects in a transgenic mouse model of Alzheimer's disease - Implications for space travel." PLoS One. 2017 Nov 29;12(11):e0186168. eCollection 2017. https://doi.org/10.1371/journal.pone.0186168 ; PubMed PMID: 29186131; PubMed Central PMCID: PMC5706673 , Nov-2017
Project Title:  Functional decline in mice with Alzheimer's-type neurodegeneration is accelerated by charge-particle radiation Reduce
Fiscal Year: FY 2014 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 02/01/2011  
End Date: 01/31/2015  
Task Last Updated: 12/04/2013 
Download report in PDF pdf
Principal Investigator/Affiliation:   Vlkolinsky, Roman  Ph.D. / Loma Linda University 
Address:  11175 Campus St 
Chan Shun Pavilion, A-1010 
Loma Linda , CA 92350-1700 
Email: rvlkolinsky@llu.edu 
Phone: 909-558-7403  
Congressional District: 41 
Web:  
Organization Type: UNIVERSITY 
Organization Name: Loma Linda University 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Nelson, Gregory  Loma Linda University 
Hartman, Richard  Ph.D. Loma Linda University 
Key Personnel Changes / Previous PI: Jerome Badaut, PhD terminated participation in our project as of July, 2013. Richard E Hartman, PhD ; Gregory Nelson, PhD ; Attila Szucs, PhD - subcontractor
Project Information: Grant/Contract No. NNX11AE41G 
Responsible Center: NASA JSC 
Grant Monitor: Simonsen, Lisa  
Center Contact:  
lisa.c.simonsen@nasa.gov 
Solicitation / Funding Source: 2010 Space Radiobiology NNJ10ZSA001N 
Grant/Contract No.: NNX11AE41G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) CNS:Risk of Acute (In-flight) and Late Central Nervous System Effects from Radiation Exposure (IRP Rev G)
Human Research Program Gaps: (1) CNS02:Does space radiation exposure elicit key events in adverse outcome pathways associated with neurological diseases? What are the key events or hallmarks, their time sequence and their associated biomarkers? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: End date is now 1/31/2015 per NSSC information (Ed., 11/5/13)

Task Description: Exposure of an astronaut’s central nervous system (CNS) to solar particle events (SPE) and galactic cosmic rays (GCR) may accelerate neurodegenerative changes and impact neuronal network activity, leading to cognitive deficits. There are similarities between radiation CNS effects and pathological processes found in the Alzheimer’s disease (AD). Common functional and structural findings include profound deficits in neuronal communication (synaptic transmission), cognitive impairments, neuro-inflammatory changes and reduced neurogenesis. These similarities lead us to hypothesize that subjects with a genetic propensity to develop AD-pathology may be excessively vulnerable to ionizing radiation. We previously showed in transgenic (TG) APP23 mice, a murine model of AD, that irradiation with 600 MeV/n iron particles accelerated the onset of electrophysiological changes in the hippocampus, a brain structure crucially involved in the formation of short-term memory. In this project we use young adult APP/PSEN1?E9 (APP/PSEN1) double transgenic (TG) mice and expose them to low doses of 150 MeV/n proton (irradiations performed at LLU proton treatment facility), 250 MeV/n silicon and 600 MeV/n iron-particle radiation to compare and quantify their detrimental effects on hippocampal functions and the onset of AD-like pathology. The APP/PSEN1 TG mice typically exhibit early-onset of age-related behavioral abnormalities and deficits in synaptic transmission. We hypothesized that exposure to even low radiation doses will accelerate the onset of age-related neurodegenerative processes, while in wild-type (WT) animals such damage may stay undetectable. Comparison of proton, silicon and iron radiation on selected neurophysiological end points in APP/PSEN1 TG mice will provide valuable information whether exposure to space radiation may exacerbate neurodegenerative processes. The functional end points (e.g. electrophysiological and behavioral changes) will be directly correlated with the expression of immunohistochemical markers of neurodegeneration, including amyloid plaque load, synaptic proteins and expression of neuroinflammatory cytokines. If such correlation is found it may indicate causative relationship between decrements in hippocampal functions and structural changes, which will help to elucidate the pathological mechanisms of radiation-induced neuronal injury and estimate the risks of low dose space radiation exposure to the CNS.

Research Impact/Earth Benefits: While the central nervous system (CNS) has been typically described as radiation-resistant tissue, we have previous electrophysiological and new behavioral evidence showing that even low doses of ionizing radiation may affect basic neuronal processes, such as synaptic transmission, neuronal excitability, and formation and consolidation of spatial memory. Specifically in the hippocampus, a brain structure intimately involved in the formation of memory, the ionizing radiation has been shown to impact synaptic excitability and plasticity. In addition, it cannot be excluded that ionizing radiation, even at very low doses of 0.1-1 Gy, may promote the onset of neurodegenerative disorders that affect the hippocampus, such as Alzheimer’s disease (AD). However, this hypothesis has not been fully tested with different low- and high-LET particles. Studying the impact of protons and high-LET radiation on neurodegenerative processes in mammalian CNS is a critical step, not only for the assessment of the space radiation risks for astronauts, but also for further development of modern cranial radiotherapies using charged particle radiation. The time-dependent changes in the CNS in patients undergoing cranial irradiations have been well documented, and they range from mild memory deficits to severe delayed demyelination and neurodegeneration. Whether low doses of charged particle radiation may accelerate the onset or affect the severity of AD-related pathology is not known. In the current project we used a murine double transgenic model of AD that we exposed to low- and high-LET charged-particle radiation to attempt to answer this question. We tested whether radiation affects the time course and severity of neurodegenerative processes in these AD-prone subjects. The combination of behavioral, electrophysiological, and histological data will help us to identify functional decrements and the neurodegenerative changes in the brains of the irradiated mice. The acquired data will improve our understanding of pathophysiological processes in irradiated and AD-affected CNS tissue.

Task Progress & Bibliography Information FY2014 
Task Progress: We received an approval for one year no-cost extension of our project ending in January 31, 2015.

Dr. Badaut, PhD, has been a co-investigator and lead for the immunohistochemical (IHC) aspect of our project. In July, Dr. Badaut had announced his intention to relocate to France and on his request his participation in our project was terminated as of July, 2013. Dr. Shalini Mehrotra, PhD, a first year postdoctoral fellow, was trained by Dr. J. Badaut. She was working with us since June 2012 and she was mostly responsible for performing the IHC analyses. Dr. Mehrotra had announced terminating her employment with LLU in August, 2013. Thus, the IHC part of our study was paused since Dr. Badaut's laboratory equipment (e.g., fluorescent microscope with Mercator software package) became unavailable. Nonetheless, we intend to complete the IHC part in full in collaboration with Dr. Nelson (co-investigator) and students trained by Dr. Badaut.

Re-allocation of funds allowed us to hire part time Mr. Gordon Harding as of September 2013, a senior research associate with significant experience in Western blotting and other protein quantification techniques. Mr. Harding performed initial experiments with presynaptic marker synaptophysin in mice irradiated with protons.

Technical progress: Summary Results by Aims.

Aim 1 & Aim 3 Activities. In accord with our statement of work (SOW), we completed all irradiations, behavioral testing, and in vitro electrophysiological experiments with protons and HZE (iron 600 MeV/n & silicon 250 MeV/n) particles. Proton irradiated APP/PSEN1 transgenic (TG) and wild-type (WT) mice were behaviorally tested pre- and 3 and 6 months post-irradiation followed by electrophysiological testing either at 6 or 9 months post-irradiation, as planned. HZE-irradiated mice (TG only) were behaviorally tested 3 and 6 months post-irradiation, followed by electrophysiological testing at 6-7 months post-irradiation. Pooling two electrophysiological time points (6 and 9 months) to one in HZE-irradiated animals was inevitable to boost the power of consequent statistical analyses. This change to the original SOW was opted for after observing considerable variability in electrophysiological results along with increased mortality specifically in iron-irradiated TG mice (likely unrelated to irradiation) that reduced the total numbers of experimental subjects per group. The change to the electrophysiological time point was applied for both HZE species to allow for direct comparisons of the radiation effects on relevant endpoints.

In total, from 2010-2013 we irradiated 78 TG and 16 WT mice with protons at Loma Linda University, Proton Treatment Facility. Twelve TG mice died; thus, electrophysiological testing was successfully performed in 82 proton-irradiated mice. We irradiated 120 TG with HZE particles at Brookhaven National Laboratories. Seven TG mice died spontaneously and thus electrophysiological testing was successfully performed in 113 animals. Ninety of these TG animals were electrophysiologically tested in 2013. We used conventional extracellular recordings to monitor both evoked synaptic responses and spontaneous activity. The behavioral and electrophysiological data from all proton- irradiated animals, including statistical evaluation, have been 95% completed. The analyses of electrophysiological and behavioral data from HZE-irradiated mice have been ~65% completed.

Behavioral analyses of proton and HZE-irradiated animals have been completed. Data from the water maze (WM) and the Barnes maze confirmed previously described deficits in spatial memory in control (0Gy) APP/PSEN1 TG mice (increased swim distance to the target area) when compared to the WT mice. We also observed that proton radiation (0.5 Gy) affected the performance of WT mice, but did not affect the performance of APP/PSEN1 TG mice. This may indicate that low radiation may not necessarily worsen the AD-like pathology, or that such pathology trumps any radiation-induced effects. In APP/PSEN1 TG mice irradiated with 600 MeV/n iron particles we surprisingly observed improved performance in WM (reduced cumulative distance to the target platform), the effect became significant at 6 months post-irradiation at the dose of 1 Gy. Interestingly, the TG mice irradiated with 250 MeV/n silicon particles exhibited reduced performance in WM at 3 months; the decrement was statistically significant at 0.1 Gy only and appeared to be transient as it could not be detected at 6 months post-irradiation. No significant differences were observed for either HZE species in the Barnes maze or zero maze.

Electrophysiological data show that proton radiation at doses from 0.1 to 1 Gy may impact synaptic excitability and short term synaptic plasticity mediated by presynaptic glutamate release, but it likely does not affect long-term potentiation (LTP; reported previously), the widely used cellular correlate of memory formation in the hippocampus. We observed that proton radiation-induced changes in synaptic excitability are qualitatively different in APP/PSEN1 TG and WT mice. In accord with our behavioral findings, the WT mice exhibit different sensitivity to radiation and, for example at 0.5 Gy we observed increased postsynaptic excitability in CA1 neurons, whereas the TG mice exhibited opposite responses at the same radiation dose. The stimulation paradigms using two (paired) stimulation pulses were used to evaluate the effect of proton radiation on presynaptic glutamate release (paired-pulse facilitation; PPF). In TG mice at 6 months post-irradiation with protons we observed reduced PPF indicating increased glutamate release and this change became more pronounced at 9 months post-irradiation. Changes in PPF were not detected in WT mice. On the other hand, WT mice exhibited sensitivity to proton radiation because at the dose of 0.5 Gy we observed radiation-induced decrements in frequency of sharp wave-ripple complexes, which are implicated in memory consolidation process in the hippocampus. Interestingly, in TG mice, a radiation exposure to protons or HZE particles had no effect on these spontaneous oscillations.

Aim 2 Activities. We partly completed immunohistological evaluations of ß-amyloid deposits in the brain samples (the cortex and the hippocampus) of APP/PSEN1 TG mice irradiated with protons using thioflavin-S staining (fibrillar form of amyloid) and by IHC using 6E10 monoclonal antibody (total amyloid). Both methods confirmed amyloid depositions in the brains of APP/PSEN1 TG mice at 6 and 9 months post irradiation. In the dorsal cortex (but not the hippocampus) at 1 Gy of protons we observed significant increase of total amyloid by 9 months post-irradiation detected by 6E10 antibodies. The IHC on brain samples irradiated with HZE particles was temporarily paused due to departure of Drs. Badaut and Methorta. Nonetheless, the IHC analyses of HZE irradiated samples is planned for the fourth year of the project (the no-cost extension has been approved) by hardware provided by Dr. Nelson (co-investigator) and performed by other team members trained in Dr. Badaut’s lab and by student volunteers.

Neuroinflammation and neurodegenerative changes in TG (and WT) brains (cortex only) exposed to radiation have been assessed by determination of five cytokines/chemokines (IL-1 beta, IL-6, TNF alpha, MCP-1, and IL-10). These molecules have been previously reported to be elevated in irradiated brains and/or have been shown to affect synaptic plasticity in the hippocampus, thus their elevation may be associated with functional decrements observed in these animals. The Luminex assays have been completed in samples irradiated with protons, the assays with HZE-irradiated brains will be completed by December, 2013. In a cohort of proton-irradiated mice we observed differences in the expression of chemokine IL-10 between TG and WT mice at 9 months, but the effect was not dependent on the radiation exposure. The other chemokines were not affected by either genotype or radiation, indicating that at 9 months radiation effects on the CNS are not associated with elevated levels of pro-inflammatory cytokines. This also indicated that the electrophysiological and behavioral decrements reported above are not due to elevated levels of cytokines within the CNS, as previously suggested by us and other investigators.

We are currently performing the analyses of synaptic markers in WT and TG mice irradiated with protons by Western blotting. The initial analyses in APP/PSEN1 TG mice irradiated with protons indicates that such exposure may increase the expression of synaptic vesicle glycoprotein and presynaptic marker synaptophysin, which may explain the radiation-induced changes in PPF described above. This marker has been previously shown to be affected by exposure to iron radiation, which awaits confirmation in APP/PSEN1 TG mice planned for the next year. Analyses in cortices irradiated with 0.1 and 1 Gy of protons and with HZE particles will be ensuing.

Bibliography Type: Description: (Last Updated: 04/24/2019) 

Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Rudobeck E, Szücs A, Vlkolinsky R. "Effects of Proton Radiation on Evoked and Spontaneous Neuronal Activity in the Hippocampus of APP/PSEN1 Transgenic Mice." HITSRS2013--Heavy Ion in Therapy and Space Radiation Symposium 2013, Chiba, Japan, May 15-18, 2013.

HITSRS2013--Heavy Ion in Therapy and Space Radiation Symposium 2013, Chiba, Japan, May 15-18, 2013. , May-2013

Abstracts for Journals and Proceedings Rudobeck E, Szücs A, Mehrotra S, Vlkolinsky R. "Ionizing radiation impairs hippocampal functions in APP/PSEN1 transgenic mice." Neuroscience 2013, San Diego, CA, November 9-13, 2013.

Neuroscience 2013, San Diego, CA, November 9-13, 2013. Program#/Poster#: 802.06/E19. Abstract available at: http://www.abstractsonline.com/Plan/ViewAbstract.aspx?sKey=f4ee62f0-ad7c-41c0-b7f1-421468a48565&cKey=24c25863-9125-4c1d-bae4-6160ce30d33f&mKey={8D2A5BEC-4825-4CD6-9439-B42BB151D1CF} ; accessed 12/5/13. , Nov-2013

Abstracts for Journals and Proceedings Bellone E, Vlkolinsky R, Hartman RE. "Low doses of iron or silicon radiation affect spatial memory in APP/PSEN1 double transgenic mice." Neuroscience 2013, San Diego, CA, November 9-13, 2013.

Neuroscience 2013, San Diego, CA, November 9-13, 2013. Program#/Poster#: 41.26/H16. Available at: http://www.abstractsonline.com/Plan/ViewAbstract.aspx?sKey=18e608b4-6987-4eaa-8a1e-448452cce869&cKey=50d60e7b-bf88-49c5-b79b-689997dbdd73&mKey={8D2A5BEC-4825-4CD6-9439-B42BB151D1CF} ; accessed 12/5/13. , Nov-2013

Articles in Peer-reviewed Journals Rudobeck E, Szücs A, Vlkolinsky R. "Effects of Proton Radiation on Evoked and Spontaneous Neuronal Activity in the Hippocampus of APP/PSEN1 Transgenic Mice." Journal of Radiation Research. In press, as of December 2013. To be published January 2014. , Dec-2013
Project Title:  Functional decline in mice with Alzheimer's-type neurodegeneration is accelerated by charge-particle radiation Reduce
Fiscal Year: FY 2013 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 02/01/2011  
End Date: 01/31/2015  
Task Last Updated: 12/04/2012 
Download report in PDF pdf
Principal Investigator/Affiliation:   Vlkolinsky, Roman  Ph.D. / Loma Linda University 
Address:  11175 Campus St 
Chan Shun Pavilion, A-1010 
Loma Linda , CA 92350-1700 
Email: rvlkolinsky@llu.edu 
Phone: 909-558-7403  
Congressional District: 41 
Web:  
Organization Type: UNIVERSITY 
Organization Name: Loma Linda University 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Nelson, Gregory  Loma Linda University 
Badaut, Jerome  Ph.D. Loma Linda University 
Hartman, Richard  Ph.D. Loma Linda University 
Key Personnel Changes / Previous PI: Jerome Badaut, PhD ; Richard E Hartman, PhD ; Gregory Nelson, PhD ; Attila Szucs, PhD - subcontractor
Project Information: Grant/Contract No. NNX11AE41G 
Responsible Center: NASA JSC 
Grant Monitor: Simonsen, Lisa  
Center Contact:  
lisa.c.simonsen@nasa.gov 
Solicitation / Funding Source: 2010 Space Radiobiology NNJ10ZSA001N 
Grant/Contract No.: NNX11AE41G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) CNS:Risk of Acute (In-flight) and Late Central Nervous System Effects from Radiation Exposure (IRP Rev G)
Human Research Program Gaps: (1) CNS02:Does space radiation exposure elicit key events in adverse outcome pathways associated with neurological diseases? What are the key events or hallmarks, their time sequence and their associated biomarkers? (IRP Rev F)
Flight Assignment/Project Notes: NOTE: End date is now 1/31/2015 per NSSC information (Ed., 11/5/13)

Task Description: Exposure of an astronaut’s central nervous system (CNS) to solar particle events (SPE) and galactic cosmic rays (GCR) may accelerate neurodegenerative changes and impact neuronal network activity, leading to cognitive deficits. There are similarities between radiation CNS effects and pathological processes found in the Alzheimer’s disease (AD). Common functional and structural findings include profound deficits in neuronal communication (synaptic transmission), cognitive impairments, neuro-inflammatory changes and reduced neurogenesis. These similarities lead us to hypothesize that subjects with a genetic propensity to develop AD-pathology may be excessively vulnerable to ionizing radiation. We previously showed in transgenic (TG) APP23 mice, a murine model of AD, that irradiation with 600 MeV/n iron particles accelerated the onset of electrophysiological changes in the hippocampus, a brain structure crucially involved in the formation of short-term memory. In this project we use young adult APP/PSEN1?E9 (APP/PS1) double transgenic (TG) mice and expose them to low doses of 150 MeV/n proton (irradiations performed at LLU proton treatment facility), 250 MeV/n silicon and 600 MeV/n iron-particle radiation to compare and quantify their detrimental effects on hippocampal functions and the onset of AD-like pathology. The APP/PS1 TG mice typically exhibit early-onset of age-related behavioral abnormalities and deficits in synaptic transmission. We hypothesized that exposure to even low radiation doses will accelerate the onset of age-related neurodegenerative processes, while in wild-type (WT) animals such damage may stay undetectable. Comparison of proton, silicon and iron radiation on selected neurophysiological end points in APP/PS1 TG mice will provide valuable information about the risks of space radiation-induced neurodegenerative processes. The functional end points (e.g. electrophysiological and behavioral changes) will be directly correlated with the expression of immunohistochemical markers of neurodegeneration, including amyloid plaque load, synaptic proteins and expression of neuroinflammatory cytokines. This information can be directly related to risks of AD onset in human subjects.

Research Impact/Earth Benefits: While the central nervous system (CNS) has been typically described as radiation-resistant tissue, we have previous electrophysiological and new behavioral evidence showing that even low doses of ionizing radiation may affect basic neuronal processes, such as synaptic transmission, neuronal excitability and formation and consolidation of spatial memory. Specifically in the hippocampus, a brain structure intimately involved in formation of memory, the ionizing radiation has been shown to impact synaptic excitability and plasticity. In addition, it cannot be excluded that ionizing radiation, even at very low doses of 0.1-1 Gy, may promote the onset of neurodegenerative disorders that affect the hippocampus, such as the Alzheimer’s disease (AD). However this hypothesis has not been fully tested with different low- and high-LET particles. Studying the impact of protons and high-LET radiation on neurodegenerative processes in mammalian CNS is a critical step, not only for the assessment of the space radiation risks for astronauts, but also for further development of modern cranial radiotherapies using charged particles. The time-dependent changes in the CNS in patients undergoing cranial irradiations have been well documented, and they range from acute mild memory deficits to severe delayed demyelination and neurodegeneration. Whether low doses of charged particle radiation may accelerate the onset or affect the severity of AD-related pathology is not known. In the current project we used a murine double transgenic model of AD that we exposed to low- and high-LET charged-particle radiation to attempt to answer this question. We test if radiation affects the time course and severity of neurodegenerative processes in these AD-prone subjects. The combination of behavioral, electrophysiological, and histological data will help us to identify functional decrements and the neurodegenerative changes in brains of the irradiated mice. The acquired data will improve our understanding of pathophysiological processes in irradiated and AD-affected CNS tissue.

Task Progress & Bibliography Information FY2013 
Task Progress: Administrative Changes: In the summer of 2012 our laboratories were relocated to new premises that was associated with remodeling, transfer, and upgrade of our electrophysiological setups and supporting apparatus. This relocation was completed in June-August 2012 and did have minor impact on our irradiation/recording/data analyses schedules. Nonetheless, several technical problems and hardware related issues needed to be solved before the electrophysiological setups became fully functional and tuned for upcoming recordings.

Technical Progress and Summary Results by Aims:

Aim 1 & Aim 3 Activities. In accord with our statement of work (SOW), we performed 7 irradiations with protons (7 irradiation runs) in APP/PS1 transgenic (TG) and wild type mice in 2011, as described in previous task book report. Irradiations were preceded with behavioral assessments (1 week prior to irradiation) that continued at 3 and 6 months post irradiation. Ensuing in vitro electrophysiological testing was performed at 6 and 9 months post irradiation with projected start in December 2011 and end in May 2012. Due to increased mortality of APP/PS1 transgenic mice irradiated with protons in summer of 2011 (12 deaths in 66 mice), we performed an additional proton irradiation run in August of 2012 using 12 APP/PS1 TG. The electrophysiological evaluation of these mice is projecting to February 2013. Their addition was required to help to identify statistical significance in cohorts with subtle changes in synaptic plasticity. The increased mortality has not been observed in any other irradiation group.

The electrophysiological experiments from the first 7 runs were completed as planned. Preliminary behavioral, electrophysiological, and histological data (see below) were presented at the NASA Space Radiation Investigators Workshop in Durham, NC, and at the Society for Neuroscience’s annual conference in New Orleans, LA, 2012 (see below). The timely analyses of the functional data (e.g. the electrophysiological and behavioral tests) in these animals was critical for directing further experiments performed in APP/PS1 transgenic mice exposed to iron and silicon radiation (irradiated at Brookhaven National Laboratories in spring, summer and fall, 2012 (Aim 2)). We used conventional extracellular recordings to monitor both evoked synaptic responses and spontaneous activity. We initially implemented the multielectrode array system (MED64, Panasonic, Japan) to record excitability at multiple neuronal fields (within each brain slice) simultaneously. However, this approach proved to be highly unreliable in APP/PS1 mice due to slice instability on the chip and its susceptibility to the outside electrical noise, which precluded reliable analyses of long-term recordings of spontaneous activity. The data analyses from conventional extracellular recordings on the short-term and long-term plasticity (long-term potentiation - LTP) have been approximately 70-80% completed. These data show a trend indicating that proton radiation may impact synaptic plasticity (LTP magnitudes) in APP/PS1 TG mice. In accord with our behavioral findings (see below), in WT mice at 0.5 Gy we observed trend showing altered (increased the magnitude) of long-term synaptic plasticity at 9 months post-irradiation. The data on excitability profiles in CA1, CA3 and DG (dentate gyrus) neurons is under evaluation. The spontaneous activity recorded in CA1 and CA3 neurons reflecting memory consolidation process in the hippocampus (and known as sharp wave-ripple complexes) required the development of specialized software (by subcontractor Dr. A. Szucz, UCSD). These data are currently being analyses.

Behavioral analyses of proton-irradiated animals have been completed. Data from the Morris Water maze (MWM) and the Barnes maze confirmed previously described deficits in spatial memory in APP/PS1 TG mice (increased swim distance to the target area) when compared to the WT mice. We also observed that proton radiation (0.5 Gy) affected performance of WT mice, but did not affect performance of APP/PS1 TG mice. This may indicate that low radiation may not necessarily worsen the AD-like pathology, or that such pathology trumps any radiation-induced effects. While histochemical and immunohistological evaluations were not planned for the 1st and 2nd year of the project, we started analysis of ß-amyloid deposits in brain samples (the cortex and the hippocampus) of APP/PS1 TG mice using thioflavin-S staining (fibrillar form of amyloid) and by IHC using 6E10 monoclonal antibody (Covance,Inc., NJ). Both methods confirmed amyloid depositions in brains of APP/PS1 TG mice at 6 and 9 months post irradiation at radiation dose of 1 Gy. However, no significant radiation-induced changes were observed. Analysis of radiation effects triggered by exposure to 0.1 and 0.5 Gy is underway.

Aim 2 & Aim 3 Activities. In accord with our SOW, we have completed all irradiation runs at Brookhaven National Laboratories. In total, we irradiated 120 APP/PS1 transgenic mice with 600 MeV/n iron and 250 MeV/n silicon particles. Radiation doses were equivalent to those in proton-irradiated mice, which will facilitate the comparisons of radiation dose effects. After completion of irradiation runs at BNL, these animals were shipped to LLU where they have been behaviorally tested at 3 and 6 months. Due to complicated legislative issues (e.g. shipping of mice from the vendor to BNL and LLU) it was not possible to test these mice behaviorally before the radiation exposure. Three and 6 month post irradiation behavioral testing was performed identically to that of mice irradiated with protons (Aim 1). Mice were then used for electrophysiological assessments of synaptic changes at 6-7 months post irradiation. Electrophysiological testing has commenced in October of 2012 and will be completed in June of 2013. At the same time, the brain tissue (left hemisphere only) of each mouse is being processed for later histochemical and immunohistological analyses planned after June of 2013.

Bibliography Type: Description: (Last Updated: 04/24/2019) 

Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Rudobeck E, Mistry N, Hartman RE, Badaut J, Vlkolinsky R. "Functional Effects of Proton Radiation on Synaptic Transmission and Plasticity in the Hippocampus of APP/PSEN1 Transgenic Mice." 23rd Annual NASA Space Radiation Investigators' Workshop, Durham, NC, July 8-11, 2012. Poster session: Poster #8084.

23rd Annual NASA Space Radiation Investigators' Workshop, Durham, NC, July 8-11, 2012. , Jul-2012

Abstracts for Journals and Proceedings Bellone JA, Hartman RE, Vlkolinsky R. "Low Doses of Proton Radiation do not Induce Spatial Learning or Memory Deficits in a Mouse Model of Alzheimer’s Disease." 23rd Annual NASA Space Radiation Investigators' Workshop, Durham, NC, July 8-11, 2012. Poster session: Poster #8004 and oral presentation.

23rd Annual NASA Space Radiation Investigators' Workshop, Durham, NC, July 8-11, 2012. , Jul-2012

Abstracts for Journals and Proceedings Bellone JA, Vlkolinsky R, Hartman RE. "The Effect of Low Doses of Proton Particle Radiation on Behavior in a Mouse Model of Alzheimer’s Disease." Society for Neuroscience 2012, New Orleans, LA, October 13-17, 2012. Poster session: Poster #343.02.

Society for Neuroscience 2012, New Orleans, LA, October 13-17, 2012. Program#/Poster#: 343.02/G11. Abstract available at: http://www.abstractsonline.com/Plan/ViewAbstract.aspx?sKey=6f2594af-aa82-405b-b329-3d7d4b117591&cKey=c34f0a5e-5059-4cd5-8935-20d2626a94ea&mKey=70007181-01c9-4de9-a0a2-eebfa14cd9f1 ; accessed 12/6/2012. , Oct-2012

Abstracts for Journals and Proceedings Mistry M, Hartman RE, Badaut J, Mehrotra S, Vlkolinsky R. "The effect of low doses of proton particle radiation on amyloid beta deposition in a mouse model of Alzheimer's disease." Society for Neuroscience 2012, New Orleans, LA, October 13-17, 2012. Posters; F30 Poster #649.18.

Society for Neuroscience 2012, New Orleans, LA, October 13-17, 2012. Program#/Poster#: 649.18/F30. Abstract available at: http://www.abstractsonline.com/Plan/ViewAbstract.aspx?sKey=72dfaad9-4c6f-4063-bb71-ac72579a2e83&cKey=b2d6efb1-a577-4e5d-9316-b8ef63155176&mKey=70007181-01c9-4de9-a0a2-eebfa14cd9f1 ; accessed 12/6/2012. , Oct-2012

Awards Bellone JA, Hartman RE, Vlkolinsky R. "John Bellone was First prize winner in student contest for 'Low Doses of Proton Radiation do not Induce Spatial Learning or Memory Deficits in a Mouse Model of Alzheimer’s Disease.' 23rd Annual NASA Space Radiation Investigators' Workshop, Durham, NC, July 8-11, 2012." Jul-2012
Awards Bellone JA, Vlkolinsky R, Hartman RE. "John Bellone was Second Prize winner for 'Effects of Proton Radiation on Behavior in a Mouse Model of Alzheimer’s Disease.' William James Excellence in Research Student Competition, LLU, October 2012." Oct-2012
Project Title:  Functional decline in mice with Alzheimer's-type neurodegeneration is accelerated by charge-particle radiation Reduce
Fiscal Year: FY 2012 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 02/01/2011  
End Date: 01/31/2014  
Task Last Updated: 12/02/2011 
Download report in PDF pdf
Principal Investigator/Affiliation:   Vlkolinsky, Roman  Ph.D. / Loma Linda University 
Address:  11175 Campus St 
Chan Shun Pavilion, A-1010 
Loma Linda , CA 92350-1700 
Email: rvlkolinsky@llu.edu 
Phone: 909-558-7403  
Congressional District: 41 
Web:  
Organization Type: UNIVERSITY 
Organization Name: Loma Linda University 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Nelson, Gregory  Loma Linda University 
Badaut, Jerome  Ph.D. Loma Linda University 
Hartman, Richard E Ph.D. Loma Linda University 
Key Personnel Changes / Previous PI: Jerome Badaut, PhD ; Richard E Hartman, PhD
Project Information: Grant/Contract No. NNX11AE41G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2010 Space Radiobiology NNJ10ZSA001N 
Grant/Contract No.: NNX11AE41G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:
No. of Master's Candidates:
No. of Bachelor's Candidates:
No. of PhD Degrees:
No. of Master's Degrees:
No. of Bachelor's Degrees:
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) CNS:Risk of Acute (In-flight) and Late Central Nervous System Effects from Radiation Exposure (IRP Rev G)
Human Research Program Gaps: (1) CNS02:Does space radiation exposure elicit key events in adverse outcome pathways associated with neurological diseases? What are the key events or hallmarks, their time sequence and their associated biomarkers? (IRP Rev F)
Task Description: Exposure of an astronaut’s central nervous system (CNS) to solar particle events (SPE) and galactic cosmic rays (GCR) may accelerate neurodegenerative changes and impact neuronal network activity leading to cognitive deficits. There are similarities between radiation CNS effects and pathological processes found in the Alzheimer’s disease (AD). Common functional and structural findings include profound deficits in neuronal communication (synaptic transmission), cognitive impairments and neuro-inflammatory changes. These similarities lead us to hypothesize that subjects with a genetic propensity to develop AD-pathology may be excessively vulnerable to ionizing radiation. We previously showed in transgenic (tg) APP23 mice, a murine model of AD, that irradiation with 600 MeV/n iron particles accelerate the onset of electrophysiological changes in the hippocampus, a brain structure crucially involved in the formation of short-term memory. In this project we use young adult APP/PSEN1?E9 (APP/PS1) double transgenic (tg) mice and expose them to low doses of 150 MeV/n proton (irradiations performed at LLU proton treatment facility), 250 MeV/n silicon and 600 MeV/n iron-particle radiation to compare and quantify their detrimental effects on hippocampal functions and onset of AD-like pathology. The APP/PS1 mice typically exhibit early-onset of age-related behavioral abnormalities and deficits in synaptic transmission. The exposure to even low radiation doses will accelerate the onset of age-related neurodegenerative processes, while in wild-type animals such damage may stay undetectable. Comparison of proton, silicon and iron radiation on selected neurophysiological end points in APP/PS1 tg mice will provide valuable information about the risks of space radiation-induced neurodegenerative processes. The functional endpoints will be directly correlated with expression of immunohistochemical markers of neurodegeneration, including amyloid plaque load, synaptic proteins and the presence of neuroinflammatory cytokines. This information can be directly related to risks of AD onset in human subjects.

Research Impact/Earth Benefits: While the central nervous system (CNS) has been typically described as radiation-resistant tissue, we have electrophysiological evidence showing that even low doses of charged-particle ionizing radiation (2-4 Gy) may affect basic neuronal processes such as synaptic transmission, neuronal excitability and formation of memory. Specifically in the hippocampus, a brain structure intimately involved in formation of memory, the ionizing radiation has been shown to be detrimental to ongoing adult neurogenesis and to synaptic plasticity. It cannot be excluded that ionizing radiation promotes the onset of neurodegenerative disorders that affect the hippocampus, such as Alzheimer’s disease (AD), however this hypothesis has not been fully tested and pathophysiological processes involved have not been characterized. In this project we use a murine double transgenic model of AD that will be exposed to charged-particle radiation. The combination of behavioral, electrophysiological, and histological data will help us to identify mechanisms of neurodegenerative changes in irradiated subjects and describe their time-course. The acquired data will not only help with assessing the radiation-related risks to astronauts, but will also improve our understanding of pathophysiological processes in the mammalian hippocampus in AD.

Task Progress & Bibliography Information FY2012 
Task Progress: The management of the grant at Loma Linda University (LLU) in a practical sense started with availability of funds at Loma Linda University on March 23, 2011. In order to successfully perform work on the project in its full scope two full-time positions were opened, for postdoctoral fellow and research technician, respectively. In addition, one pre-doctoral student has been involved in behavioral testing.

Irradiations: From May to August 2011 we have irradiated 82 mice (66 transgenic (tg) APP/PS1 and 16 wild-type (wt) mice) with proton beam (150 MeV/n) at the age of 3 mo. Irradiations were performed according to the plan in 7 batches that were separated by 1-2 weeks. This time separation was important for the upcoming use of animals for electrophysiological experiments (6 and 9 mo later) with their rate-limiting step of 1 animal/day. Each batch contained 10-12 mice that were irradiated with 0, 0.1, 0.5 and 1.0 Gy (whole body) and they were ascribed to either 6 or 9 mo age groups. The first electrophysiological experiments will commence on December 6th, 2011. The first irradiation with iron 600 MeV/n at Brookhaven National Laboratories is scheduled in spring and summer 2012.

Animal mortality: We observed increased mortality in the 1st batch of tg mice likely unrelated to irradiation, but possibly due to AD-like pathology. We plan on adding more subjects to the experimental groups to maintain the statistical power. However, if overall mortality in all cohort of animals, and specifically in batch 1 at 9 mo post-irradiation increases above 25-30%, we may need to shorten the post-irradiation interval for whole experiment from 9 mo to 6 mo. The decision will be made when first electrophysiological and histological data (specifically the thio-S staining of amyloid plaque load) become available (spring 2012) and AD-like pathology at 6 mo. will be confirmed.

Behavioral testing before irradiation: In addition to our original proposal we decided to establish pre-irradiation baseline values for hippocampus-dependent behavior using the Morris Water Maze (MWM). This additional time point will promote paired-comparisons pre- vs. post-irradiation for each animal. Pre-irradiation data were recorded within 1 week after delivery of the animals to LLU. First, cued variant of MWM has been used to exclude differences in swimming ability, visual deficits, and/or motivation between APP/PS1 tg and wild-type (wt) mice, which was prerequisite for further successful behavioral testing. Next, hippocampus-dependent spatial learning (two spatial tests) was assessed in 2 consecutive days showing that wt mice performed better in spatial tasks. However, in wt mice we also observed unexpected freezing behavior that typically reflects an increased anxiety. These data require further evaluation. Up to date, all animals were tested at 3 mo. post-irradiation and batch 1 is currently undergoing behavioral testing at 6 mo post-irradiation, which will be followed with in vitro electrophysiology (Dec 6th, 2011). In addition to MWM, at 3 and 6 mo. post-irradiation, we have also included the Barnes Maze test that should strengthen our findings on spatial learning and memory deficits between irradiated groups.

Technical Highlights: A new MED64 multi-electrode array setup has been assembled and will be used for measurements of spontaneous synaptic activity in the hippocampus of APP/PS1 tg mice. It has been tuned to record network activity in brain slices in vitro, in all hippocampal fields simultaneously using P5001A probes (MED64, Panasonic, Japan). At the same time conventional recordings will be performed using glass microelectrodes to assess radiation-induced changes in slice excitability that cannot be fully examined by MED64 due to limitations in stimulation currents. Our effort is currently directed towards electrophysiological recordings of spontaneous network activity, where spontaneous activity is promoted either by pharmacological manipulation of excitatory vs. inhibitory drive (e.g. blockage of GABA-ergic interneurons by bicuculline, activation of NMDA receptors by lowered extracellular magnesium or by stimulation of cholinergic inputs by physostigmine), or by electrical stimulation of afferent pathways and induction of long-term potentiation (LTP) of synaptic transmission. All these manipulations are known to promote spontaneous activity in the hippocampus and are relevant to pathological processes found in AD. These electrophysiological experiments are at their introductory phase. The first complete set of data acquired from APP/PS1 tg mice is expected to be available at the end of December 2011.

Bibliography Type: Description: (Last Updated: 04/24/2019) 

Show Cumulative Bibliography Listing
 
 None in FY 2012
Project Title:  Functional decline in mice with Alzheimer's-type neurodegeneration is accelerated by charge-particle radiation Reduce
Fiscal Year: FY 2011 
Division: Human Research 
Research Discipline/Element:
HRP SR:Space Radiation
Start Date: 02/01/2011  
End Date: 01/31/2014  
Task Last Updated: 03/16/2011 
Download report in PDF pdf
Principal Investigator/Affiliation:   Vlkolinsky, Roman  Ph.D. / Loma Linda University 
Address:  11175 Campus St 
Chan Shun Pavilion, A-1010 
Loma Linda , CA 92350-1700 
Email: rvlkolinsky@llu.edu 
Phone: 909-558-7403  
Congressional District: 41 
Web:  
Organization Type: UNIVERSITY 
Organization Name: Loma Linda University 
Joint Agency:  
Comments:  
Co-Investigator(s)
Affiliation: 
Nelson, Gregory  Loma Linda University 
Project Information: Grant/Contract No. NNX11AE41G 
Responsible Center: NASA JSC 
Grant Monitor: Cucinott1a, Francis  
Center Contact: 281-483-0968 
noaccess@nasa.gov 
Solicitation / Funding Source: 2010 Space Radiobiology NNJ10ZSA001N 
Grant/Contract No.: NNX11AE41G 
Project Type: GROUND 
Flight Program:  
TechPort: No 
No. of Post Docs:  
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:  
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Human Research Program Elements: (1) SR:Space Radiation
Human Research Program Risks: (1) CNS:Risk of Acute (In-flight) and Late Central Nervous System Effects from Radiation Exposure (IRP Rev G)
Human Research Program Gaps: (1) CNS02:Does space radiation exposure elicit key events in adverse outcome pathways associated with neurological diseases? What are the key events or hallmarks, their time sequence and their associated biomarkers? (IRP Rev F)
Task Description: An unavoidable complication of space travel is exposure to proton and high-charge, high-energy (HZE) particle radiation. HZE radiation triggers oxidative stress, neuroinflammation and synaptic changes in the CNS that are reminiscent of those seen in aging or Alzheimer's disease (AD). We showed that irradiation with 56Fe particles in young adult mice accelerates the onset of electrophysiological decrements observed as reduced synaptic excitability in the hippocampus. The APP/PSEN1 double transgenic (tg) mouse (commercially available mouse model of AD) exhibits age-related behavioral abnormalities and deficits in synaptic transmission. We propose to expose young adult APP/PSEN1 tg mice to low doses of proton, helium, silicon and iron-particle radiation (brain-only) to quantify their detrimental effects on hippocampal functions. The effects of proton, helium and silicon-particle radiation on neurodegenerative processes in the CNS have not been tested. Moreover, we will compare single and fractionated exposures that may activate compensatory protective mechanisms in the neuronal tissue and significantly modify the pathophysiological process. We will record synaptic transmission in the hippocampus at 3 months (1 month after irradiation) and then at the ages of 6, 9 and 12 months. We will use multielectrode array system (MED64) that allows electrophysiological recordings of excitatory synaptic transmission, neuronal excitability and spontaneous network activity in isolated hippocampal slices in all major neuronal fields simultaneously. These electrophysiological parameters reflect the functional status of the neuronal tissue exposed to radiation and can be extrapolated to the in vivo hippocampus. The functional endpoints will be directly correlated with expression of immunohistochemical markers of neurodegeneration, including amyloid plaque load, synaptic proteins and the presence of neuroinflammatory cytokines.

Research Impact/Earth Benefits: 0

Task Progress & Bibliography Information FY2011 
Task Progress: New project for FY2011.

Bibliography Type: Description: (Last Updated: 04/24/2019) 

Show Cumulative Bibliography Listing
 
 None in FY 2011