Combination of Mn²⁺‐enhanced and diffusion tensor MR imaging gives complementary information about injury and regeneration in the adult rat optic nerve

Abstract

Purpose:To evaluate manganese (Mn 2ϩ )-enhanced MRI (MEMRI) and diffusion tensor imaging (DTI) as tools for detection of axonal injury and regeneration after intravitreal peripheral nerve graft (PNG) implantation in the rat optic nerve (ON). Materials and Methods:In adult Fischer rats, retinal ganglion cell (RGC) survival was evaluated in Flurogold (FG) back-filled retinal whole mounts after ON crush (ONC), intravitreal PNG, and intravitreal MnCl 2 injection (150 nmol) at 0 and 20 days post lesion (dpl). MEMRI and echoplanar DTI (DTI-EPI) was obtained of noninjured ON one day after intravitreal MnCl 2 injection, and at 1 and 21 dpl after ONC, intravitreal PNG, and intravitreal MnCl 2 injections given at 0 and 20 dpl. GAP-43 immunohistochemistry was performed after the last MRI.Results: ONC reduced RGC density in retina by 94% at 21 dpl compared to noninjured ON without MnCl 2 injections. Both intravitreal PNG and intravitreal MnCl 2 injections improved RGC survival in retina, which was reduced by 90% (ONCϩMnCl 2 ), 82% (ONCϩPNG), and 74% (ONCϩPNGϩMnCl 2 ) compared to noninjured ON. DTIderived parameters (fractional anisotropy [FA], mean diffusivity, axial diffusivity , and radial diffusivity Ќ ) were unaffected by the presence of Mn 2ϩ in the ON. At 1 dpl, CNR MEMRI and were reduced at the injury site, while at 21 dpl they were increased at the injury site compared to values measured at 1 dpl. GAP-43 immunoreactive axons were present in the ON distal to the ONC injury site. Conclusion:MEMRI and DTI enabled detection of functional and structural degradation after rat ON injury, and there was correlation between the MRI-derived and immunohistochemical measures of axon regeneration. UNLIKE AXONS IN THE PERIPHERAL NERVOUS SYS-TEM, those in the central nervous system (CNS) of adult mammals do not regenerate after injury (1,2). Failure to regenerate is attributed to a combination of axon growth arrest by myelin-associated and scar-derived inhibitory molecules (3,4), and to the absence of growth-promoting neurotrophic factors in the adult CNS (5). Several therapeutic interventions have been tested in animal models to try to promote axon regeneration in the adult mammalian CNS, including neutralizing inhibitory molecules by bacterial enzyme chondroitinase ABC (6) and administration of growthpromoting factors released, for example, from olfactory ensheathing cells and stem cells (7,8). One method that has been shown to have an effect in an optic nerve (ON) animal model is the intravitreal implantation of a peripheral nerve graft (PNG) after ON transection. Schwann cells in the PNG produce trophic factors that promote both retinal ganglion cell (RGC) survival and axon growth through the putative inhibitory environment of the injured ON, as documented in several studies (9 -11). Most studies detect regenerating axons in the CNS using traditional axon tracing techniques (e.g.,

MR Imaging of Papilledema and Visual Pathways: Effects of Increased Intracranial Pressure and Pathophysiologic Mechanisms

“…61 Other experiments in rats have demonstrated decreased anisotropy within the ON following glaucoma induction and ON injury. 54,62 DTI has demonstrated decreased anisotropy and increased diffusivity in children with ON hypoplasia in septooptic dysplasia. 63 Disruption of these pathways leading to atrophy of distal axons projecting from the retina 64 could predict visual loss from degeneration related to papilledema.…”

“…52 While not applicable to human studies, manganese-enhanced MR imaging may give additional insight into ON function by using the property of manganese uptake by active neurons to image neural activity. [53][54][55][56][57] Visualization of the fiber tracts from the retina to the visual cortex is also being investigated with DTI. 58,59 DTI can map optic radiations within the brain with a great deal of accuracy compared with postmortem dissections.…”

Axonal tracing of the normal and regenerating visual pathway of mouse, rat, frog, and fish using manganese‐enhanced MRI (MEMRI)

“…Here, we investigate the feasibility of manganeseenhanced MRI (MEMRI) as a possible longitudinal axon tracing method for intra-orbitally transected ON axons spontaneously regenerating in frogs and fish, and compare our findings with MEMRI of axons induced to regenerate in adult rats and mice after intravitreal grafting of peripheral nerve implants and lens injury, respectively. MEMRI has been developed by us (12)(13)(14)(15) and others (16)(17)(18) In the present study, we test the hypothesis that MEMRI has the potential to longitudinally image the 100% of spontaneously regenerating axons in frogs and fish by comparing MEMRI of regenerating ON in these species with the comparatively moderate regrowth of axons induced in rats by intravitreal PNS implants and in mice by LI. We predict that, if the limitations of MEMRI are equally applicable to the imaging of normal and regenerating fish/frog axons, similar MRI signal strengths will be recorded from normal and regenerating ON, because the densities of Mn 2þ -filled axons will be similar in each group.…”

Complementary research models and methods to study axonal regeneration in the vertebrate retinofugal system

“…This technique makes use of the paramagnetic properties of manganese ions (Mn 2+ ), which are taken up by neurons via L-type voltage-gated Ca 2+ channels (Narita et al 1990), enabling the imaging of regenerating RGC axons (Ryu et al 2002;Thuen et al 2005Thuen et al , 2009Chan et al 2008;Haenold et al 2012;Sandvig et al 2012;Fischer et al 2014;Sandvig and Sandvig 2014;Yang et al 2016). To visualize optic nerve regeneration, MnCl 2 is injected intravitreally, where it is taken up by the RGCs and anterogradely transported along axonal microtubule.…”