Strategies to induce recovery from lesions of the spinal cord have not fully resulted in clinical applications. This is a consequence of a number of impediments that axons encounter when trying to regrow beyond the lesion site, and that intraspinal rearrangements are subjected to. In the present study we evaluated (1) the possibility to improve locomotor recovery after complete transection of the spinal cord by means of an adeno-associated (AAV) viral vector expressing the neurotrophin brain-derived neurotrophic factor (BDNF) in lumbar spinal neurons caudal to the lesion site and (2) how the spinal cord transection and BDNF treatment affected neurotransmission in the segments caudal to the lesion site. BDNF overexpression resulted in clear increases in expression levels of molecules involved in glutamatergic (VGluT2) and GABAergic (GABA, GAD65, GAD67) neurotransmission in parallel with a reduction of the potassium-chloride co-transporter (KCC2) which contributes to an inhibitory neurotransmission. BDNF treated animals showed significant improvements in assisted locomotor performance, and performed locomotor movements with body weight support and plantar foot placement on a moving treadmill. These positive effects of BDNF local overexpression were detectable as early as two weeks after spinal cord transection and viral vector application and lasted for at least 7 weeks. Gradually increasing frequencies of clonic movements at the end of the experiment attenuated the quality of treadmill walking. These data indicate that BDNF has the potential to enhance the functionality of isolated lumbar circuits, but also that BDNF levels have to be tightly controlled to prevent hyperexcitability.

Previous evidence indicates that locomotor exercise is a powerful means of increasing brain-derived neurotrophic factor (BDNF) and its signal transduction receptor TrkB mRNA levels, immunolabeling intensity and number of BDNF- and TrkB-immunopositive cells in the spinal cord of adult rats but the contribution of specific cell types to changes resulting from long-term activity is unknown. As changes in BDNF protein distribution due to systemic stimuli may reflect either its in-situ synthesis or its translocation from other sources, we investigated where BDNF and TrkB mRNA are expressed in the spinal lumbar segments. We report on the cell types defined by size, BDNF mRNA levels and number of cells with TrkB transcripts in sedentary and exercised animals following 28 days of treadmill walking. In the majority of cells, exercise increased perikaryonal levels of BDNF mRNA but did not affect TrkB transcript levels. Bidirectional changes in a number of TrkB mRNA-expressing cells occurred in small groups of ventral horn neurons. An increase in BDNF transcripts was translated into changes in pro-BDNF and BDNF levels. A 7-day walking regimen increased BDNF protein levels similarly to 28-day treadmill walking. Our observations indicate that long- and short-term locomotor activity of moderate intensity produce stimuli sufficient to recruit a majority of spinal cells to increased BDNF synthesis, suggesting that continuous tuning of pro-BDNF and BDNF levels permits spinal networks to undergo trophic modulation not requiring changes in TrkB mRNA supply.

The importance of neurotrophin 3 (NT-3) for motor control prompted us to ask the question whether direct electrical stimulation of low-threshold muscle afferents, strengthening the proprioceptive signaling, could effectively increase the endogenous pool of this neurotrophin and its receptor TrkC in the Hoffmann-reflex (H-reflex) circuitry. The effects were compared with those of brain-derived neurotrophic factor (BDNF) and its TrkB receptor. Continuous bursts of stimuli were delivered unilaterally for seven days, 80 min daily, by means of a cuff-electrode implanted over the tibial nerve in awake rats. The H-reflex was recorded in the soleus muscle to control the strength of stimulation. Stimulation aimed at activation of Ia fibers produced a strong increase of NT-3 protein, measured with ELISA, in the lumbar L3-6 segments of the spinal cord and in the soleus muscle. This stimulation exerted much weaker effect on BDNF protein level which slightly increased only in L3-6 segments of the spinal cord. Increased protein level of NT-3 and BDNF corresponded to the changes of NT-3 mRNA and BDNF mRNA expression in L3-6 segments but not in the soleus muscle. We disclosed tissue-specificity of TrkC mRNA and TrkB mRNA responses. In the spinal cord TrkC and TrkB transcripts tended to decrease, whereas in the soleus muscle TrkB mRNA decreased and TrkC mRNA expression strongly increased, suggesting that stimulation of Ia fibers leads to sensitization of the soleus muscle to NT-3 signaling. The possibility of increasing NT-3/TrkC signaling in the neuromuscular system, with minor effects on BDNF/TrkB signaling, by means of low-threshold electrical stimulation of peripheral nerves, which in humans might be applied in non-invasive way, offers an attractive therapeutic tool.

Bacterial lipopolysaccharide activates Toll-like receptor 4 (TLR4) and triggers proinflammatory reactions of macrophages. TLR4 signaling is negatively regulated by Lyn tyrosine kinase, provided the kinase accumulates in membrane rafts as a result of palmitoylation, the catalytic activity, and SH2- and SH3-mediated intermolecular interactions.