P2‐011: Using DTI for non‐invasive evaluation of axonal damage caused by exposure of axon terminals to amyloid beta

Background: Amyloid Beta (AB) is the major toxic component in the brains f Alzheimer's disease (AD). However, the mechanism of how AB induces neural damage remains not clear. The progression of AD begins with the synaptic eficits leading to memory loss. In line with this symptom, the accumulatedABin synaptic areas have found to depress various synaptic activities via egulating several lines of serine/threonine kinases pathways.However, synapses have high plasticity.What is more irreversible is the damage to the axons nd neuronal bodies. Although many studies have shown close correlations of axonal transport deficits and the presence of AB, these studies could not tell hether the axonal terminal AB or soma AB are essential for such a axonal impairment. Since the axonal terminal accumulated AB has shown to early ynaptic damage inAD, it is critical to knowwhether axonal terminal exposure toAB(but not somaAB) is sufficient to cause axonal damage. In this study,we sed a novel imaging technique, the Magnetic Resonance Diffusion Tensor Imaging (DTI), to non-invasively evaluate axonal degeneration caused by he axonal terminal treated with AB1-42. Our previous studies has demonstrated that high resolution DTI is capable to detect white matter damage involved n AD transgenic mice and animal models of human Multiple Sclerosis. Using axial and radial diffusivities calculated from DTI can characterize he process of axonal degeneration in optic nerves and optic tracts as a result of retinal ganglion cell ischemic damage.One unique feature of the visual ystemof the optic nerve and optic tract is that the axons are originated retinal (Graph presented) ganglion cells (RGCs). Thus, the axonal terminals of RGC locate inside the rain, while their cell bodies (soma) are outside of the brain (in the ocular space). Because of this unique anatomical feature, AB, injected directly to he optic tract axonal terminals can only affect the axon but not the soma of RGC.UsingDiffusion Tensor Imaging (DTI) and histology,we asked whether xonal exposure toABcould cause damageswithout first affecting their soma. Method(s): Ten female C57BL/6 mice at 12 weeks old were anesthetized by .5% isoflurane/oxygen using an isoflurane vaporizer (VetEquip, Pleasanton, CA).HumanAB1-42 (4 nmole,A9810, SigmaAldrich,USA)were injected at volume of 3 ul into the right hemisphere optic tract axonal terminals (coordinates: posterior 2.0 mm from the bregma, lateral 2.0 mm, ventrally 2.5 mm (Figure presented) rom the cortical surface). In 1 and 3 months after AB injection, mice were placed in holders to immobilize the heads. A 7-cm inner volume coil was sed as a transmitter coil and a 1.5-cm inner diameter surface coil was used as a receiver to collect data in aBruker 4.7TBioSpec small animalMRI instrument. mageswith slice thickness 0.5mm, field of viewof 2cmx 2cmandmatrix 128 x 128 (zero filling to 256 x 256) was collected to cover the visual ystemfrom eyes to Superior Colliculus. Spin-echo Diffusion Tensor Imaging (DTI)was performedwithTR3 s, TE29 ms, duration between a diffusion gradient air = 20 ms, diffusion gradient duration = 3 ms, and six-direction diffusion schemewith b-values of 0 and 0.85 ms/mm2. Using software written in atlab (MathWorks, Natick, MA, USA), the eigenvalues derived from diffusion tensor were used to calculate axial diffusivity, radial diffusivity, relative nisotropy (RA), and trace of the diffusion tensor (Tr). To evaluate the axonal damages in optic nerves and optic tracts, animals were sacrificed in 3 months fter AB treatment. The integrity of axons was evaluated using a primary antibody against phosphorylated neurofilament (pNF, SMI-31), and myelin integrity as assessed with a primary antibody against myelin basic protein (MBP). Histological sections were examined using an Olympus Fluoview onfocal Microscope equipped with a 60x oil objective for further analysis. Result(s): Typical DTI maps from normal and AB-treated mice were shown n Figure 1, in which the optic nerves and tracts were pointed by arrows. Among the measured white matter tracts, only optic tracts, and optic nerves howed significantDTI changes caused byAB. In optic tract, the right side (ipsilateral side) of the tracts showed a 12-16% decrease of axial diffusivity in 1 nd 3 months after AB injection while the left side of the tracts remained normal (Figure 2). As for optic nerves significant changeswere found in Trmaps, hich appeared a 13%reduction in the left nerves but appeared normal in the right nerves (Figure 3). Such a change of Tr in the left nerves closely related to he decrease of axial but the change of axial diffusivity did not reach a significant level. Immunohistochemistry showed results inconsistent with the DTI indings. SMI-31 staining was used to detect axonal integrity, and MBP staining was used to characterizemyelin loss. Comparing the left and right optic ract (Figure 4), right optic tract showed severe loss of axons (Figure 5). Conclusion(s): The importance of this study can be summarized in two aspects. irst, this is the first study that demonstrated a potential pathway that axonal damage which can be induced by and exposure of axonal terminal to AB without irst affecting the neuronal cell bodies.We used the unique anatomical feature of retinal ganglion cells with their cell bodies in the eye but the elongated xons reaching to themiddle of the brain. Micro-injection ofAB1-42 in the axonal terminals of optic tract caused optic tract axonal damage in 1 month (based n DTI). Three months after AB treatment, the histological examination showed severe axonal loss in ipsilateral optic tracts (Figure 4). Given the monocular ature of the mouse visual pathway, i.e. 96% RGC axons projecting to the opposite hemisphere, the damage to the ipsilateral optic tract would cause ore damage in the contralateral than the ipsilateral optic nerves. In line with our prediction, severe damage was seen in the contralateral but not ipsilateral (Figure presented) ptic nerves (Figure 5). Using the mouse visual system, we demonstrated a potential mechanism of axonal degeneration of the central nervous system in the rains of AD. Another important finding of this study is to demonstrate the feasibility of using noninvasive DTI imaging markers to evaluate axonal damage rogress inAD.DTI is a powerfulMRI imaging modal,which is very sensitive to detect changes of tissue microstructure in white matter. Studies have found hanges of DTI in patients in association with cognitive impairment. DTI is a clinical available imaging modality. This finding has significant clinical impact s to use of DTI for diagnosis of AD and evaluation of the treatment.