Acute mitochondrial damage and the energy crisis following axonal injury highlight mitochondrial transport as an important target for axonal regeneration. Syntaphilin (Snph), known for its potent mitochondrial anchoring action, has emerged as a significant inhibitor of both mitochondrial transport and axonal regeneration. Therefore, investigating the molecular mechanisms that influence the expression levels of the snph gene can provide a viable strategy to regulate mitochondrial trafficking and enhance axonal regeneration. Here, we reveal the inhibitory effect of microRNA-146b (miR-146b) on the expression of the homologous zebrafish gene syntaphilin b (snphb). Through CRISPR/Cas9 and single-cell electroporation, we elucidated the positive regulatory effect of the miR-146b-snphb axis on Mauthner cell (M-cell) axon regeneration at the global and single-cell levels. Through escape response tests, we show that miR-146b-snphb signaling positively regulates functional recovery after M-cell axon injury. In addition, continuous dynamic imaging in vivo showed that reprogramming miR-146b significantly promotes axonal mitochondrial trafficking in the pre-injury and early stages of regeneration. Our study reveals an intrinsic axonal regeneration regulatory axis that promotes axonal regeneration by reprogramming mitochondrial transport and anchoring. This regulation involves noncoding RNA, and mitochondria-associated genes may provide a potential opportunity for the repair of central nervous system injury.
Animals ; Zebrafish ; MicroRNAs/genetics* ; Nerve Regeneration/physiology* ; Mitochondria/metabolism* ; Zebrafish Proteins/genetics* ; Axons/metabolism* ; Signal Transduction/physiology* ; Axonal Transport/physiology* ; Nerve Tissue Proteins/genetics*

Axonal transport of mitochondria is critical for neuronal survival and function. Automatically quantifying and analyzing mitochondrial movement in a large quantity remain challenging. Here, we report an efficient method for imaging and quantifying axonal mitochondrial transport using microfluidic-chamber-cultured neurons together with a newly developed analysis package named "MitoQuant". This tool-kit consists of an automated program for tracking mitochondrial movement inside live neuronal axons and a transient-velocity analysis program for analyzing dynamic movement patterns of mitochondria. Using this method, we examined axonal mitochondrial movement both in cultured mammalian neurons and in motor neuron axons of Drosophila in vivo. In 3 different paradigms (temperature changes, drug treatment and genetic manipulation) that affect mitochondria, we have shown that this new method is highly efficient and sensitive for detecting changes in mitochondrial movement. The method significantly enhanced our ability to quantitatively analyze axonal mitochondrial movement and allowed us to detect dynamic changes in axonal mitochondrial transport that were not detected by traditional kymographic analyses.
Animals ; Axonal Transport ; physiology ; Cerebral Cortex ; cytology ; metabolism ; Drosophila melanogaster ; cytology ; metabolism ; Embryo, Mammalian ; Gene Expression ; Lab-On-A-Chip Devices ; Microscopy, Confocal ; Mitochondria ; metabolism ; ultrastructure ; Motor Neurons ; metabolism ; ultrastructure ; Movement ; Mutation ; Primary Cell Culture ; RNA-Binding Protein FUS ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Software

OBJECTIVETo study the changes in microtubule motor protein expression in the spinal cord and sciatic nerve of rats exposed to carbon disulfide, and to investigate the possible molecular mechanism of changes in axonal transport in carbon disulfide-induced peripheral neuropathy.

METHODSHealthy adult male Wistar rats were randomly divided into one control group and three experimental groups (10 rats per group). The rats in experimental groups were intoxicated by gavage of carbon disulfide at a dose of 200, 400, or 600 mg/kg 6 times a week for 6 consecutive weeks, while the rats in control group were given the same volume of corn oil by gavage. Animals were sacrificed after exposure, with nerve tissue separated. The levels of dynein, dynactin, and kinesin in the spinal cord and sciatic nerve were determined by Western blot.

RESULTSThe content of dynein, dynactin, and kinesin in the sciatic nerve decreased significantly under exposure to carbon disulfide. The levels of dynein in the sciatic nerve were reduced by 23.47% and 33.34% at exposure doses of 400 and 600 mg/kg, respectively. The levels of dynactin in the sciatic nerve of the three experimental groups were reduced by 19.91%, 24.23%, and 41.30%, respectively. The level of kinesin was reduced by 25.98%under exposure to 600 mg/kg carbon disulfide. All the differences were statistically significant (P < 0.01). As compared with the control group, the 600 mg/kg group experienced a 28.24% decrease in level of dynactin in the spinal cord (P < 0.01), but no significant change was observed in the level of dynein or kinesin.

CONCLUSIONCarbon disulfide has an impact on microtubule motor protein expression in nerve tissues, which might be involved in the development of carbon disulfide-induced peripheral neuropathy.

Animals ; Axonal Transport ; drug effects ; physiology ; Carbon Disulfide ; toxicity ; Dynactin Complex ; Male ; Microtubule-Associated Proteins ; metabolism ; Nerve Tissue ; metabolism ; Peripheral Nervous System Diseases ; chemically induced ; metabolism ; Rats, Wistar ; Sciatic Nerve ; metabolism ; Spinal Cord ; metabolism

OBJECTIVETo review the updated research on neuroprotection in glaucoma, and summarize the potential agents investigated so far.

DATA SOURCESThe data in this review were collected from PubMed and Google Scholar databases published in English up to September 2012, with keywords including glaucoma, neuroprotection, and retinal ganglion cells, both alone and in combination. Publications from the past ten years were selected, but important older articles were not excluded.

STUDY SELECTIONArticles about neuroprotection in glaucoma were selected and reviewed, and those that are cited in articles identified by this search strategy and judged relevant to this review were also included.

RESULTSAlthough lowering the intraocular pressure is the only therapy approved as being effective in the treatment of glaucoma, increasing numbers of studies have discovered various mechanisms of retinal ganglion cells death in the glaucoma and relevant neuroprotective strategies. These strategies target neurotrophic factor deprivation, excitotoxic damage, oxidative stress, mitochondrial dysfunction, inflammation, activation of intrinsic and extrinsic apoptotic signals, ischemia, and protein misfolding. Exploring the mechanism of axonal transport failure, synaptic dysfunction, the glial system in glaucoma, and stem cell used in glaucoma constitute promising research areas of the future.

CONCLUSIONSNeuroprotective strategies continue to be refined, and future deep investment in researching the pathogenesis of glaucoma may provide novel and practical neuroprotection tactics. Establishing a system to assess the effects of neuroprotection treatments may further facilitate this research.

Apoptosis ; Axonal Transport ; Brain-Derived Neurotrophic Factor ; physiology ; Ciliary Neurotrophic Factor ; physiology ; Glaucoma ; etiology ; therapy ; Humans ; Mitochondria ; physiology ; Neuroprotective Agents ; therapeutic use ; Oxidative Stress ; Protein Folding ; Receptors, N-Methyl-D-Aspartate ; physiology ; Retinal Ganglion Cells ; physiology

OBJECTIVETo investigate in vivo survival of retinal ganglion cells (RGCs) after partial blockage of optic nerve (ON) axoplasmic flow by sub-retinal space or vitreous cavity injection of brain-derived neural factor (BDNF) produced by genetically modified neural progenitor cells (NPCs).

METHODSAdult Sprague-Dawley (SD) rat RGCs were labeled with granular blue (GB) applied to their main targets in the brain. Seven days later, the left ON was intra-obitally crushed with a 40 g power forceps to partially block ON axoplasmic flow. Animals were randomized to three groups. The left eye of each rat received a sham injection, NPCs injection or an injection of genetically modified neural progenitors producing BDNF (BDNF-NPCs). Seven, 15 and 30 days after ON crush, retinas were examined under a fluorescence microscope. By calculating and comparing the average RGCs densities and RGC apoptosis density, RGC survival was estimated and the neuro-protective effect of transplanted cells was evaluated.

RESULTSSeven, 15 and 30 days after crush, in the intra-vitreous injection group, mean RGC densities had decreased to 1885 +/- 68, 1562 +/- 20, 1380 +/- 7 and 1837 +/- 46, 1561 +/- 58, 1370 +/- 16, respectively with sham injection or neural progenitors injection. However, RGCs density in the groups treated with intra-vitreous injection of BDNF-NPC was 2101 +/- 15, 1809 +/- 19 and 1625 +/- 34. Similar results were found in groups after sub-retinal injection. Higher densities were observed in groups treated with BDNF-NPCs. There were statistically significant differences among groups through nonparametric tests followed by the Mann-Whitely test. RGC apoptosis density in BDNF-NPC at each follow-up time was less than in other groups.

CONCLUSIONSA continuous supply of neurotrophic factors by the injection of genetically modified neural progenitors presents a highly effective approach to counteract optic neuropathy and RGC degeneration after partial ON axoplasmic flow blockage.

Animals ; Apoptosis ; Axonal Transport ; Brain-Derived Neurotrophic Factor ; genetics ; Cell Survival ; Gene Transfer Techniques ; Genetic Therapy ; Glaucoma ; therapy ; Male ; Rats ; Rats, Sprague-Dawley ; Retinal Ganglion Cells ; cytology ; Stem Cells ; physiology ; Vitreous Body ; metabolism

Time-dependent translocational changes of Synapsin I (SyI), a synaptic vesicle-associated phosphoprotein and its involvement in the axonal transport were investigated in the regenerating axonal sprouts. A weak SyI immunoreactivity (IR) was found in the axoplasm of normal axons. Rat sciatic nerves were crush-injured by ligating with 1-0 silk thread at the mid-thigh level and released from the ligation 24 h later. At various times after release, immunocytochemistry was performed. SyI was translocated from the proximal to the distal site of ligation and also involved in the sprouting of regenerating axons. The distribution patterns of SyI IR were changed in the crush-injured nerves. SyI immunoreactive thin processes were strongly appeared in the proximal region from 1 h after release. After 3 h, a very strong IR was expressed. The intense SyI immunoreactive thin processes were elongated distally and were changed the distribution pattern by time-lapse. After 12 h, strong immunoreactive processes were extended to the ligation crush site. At 1 day, a very intense IR was expressed. At 2 days, immunoreactive thin processes extended into the distal region over the ligation crush site and strong IR was observed after 3 days. SyI was accumulated in the proximal region at the early phases after release. These results suggest that SyI may be related to the translocation of vesicles to the elongated membranes by a fast axonal transport in the regenerating sprouts.
Animals ; Axonal Transport ; Axons/*physiology/ultrastructure ; Immunohistochemistry ; Male ; Nerve Crush ; Nerve Regeneration/*physiology ; Protein Transport ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve/physiology ; Synapsins/*metabolism ; Time Factors