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1

The effect of high-frequency, low-threshold electrical stimulation of peripheral nerves on the pool of neurotrophin 3 in the lumbar spinal cord and soleus muscle of the rat

100%
Gajewska O., Piasecka J., Kasicki S., Skup M., Czarkowska-Bauch J.
Acta Neurobiologiae Experimentalis
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2011
|
tom 71
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nr S
Locomotor exercise, sufficient to increase expression of brainderived neurotrophic factor and neurotrophin 4 in the lumbar spinal cord, does not affect protein level of neurotrophin 3 (NT-3), as we have shown previously. The effect of 7 days of direct, lowfrequency electrical stimulation of the tibial nerve on expression of NT-3 was also negligible although this stimulation was addressed to low-threshold muscle afferents expressing the NT-3 and its high-affinity receptor trkC. To verify whether upregulation of NT-3 requires stronger stimulation, we maximized efficiency of electrical stimulation. Hoffmann reflex, recorded from the soleus muscle, allowed controlling low-threshold stimulation delivered by cuff electrode implanted over the tibial nerve. Electrodes were implanted bilaterally. The nerve was stimulated unilaterally for 7 days, starting 3 weeks after surgery. The contralateral limb served as a control. Series of 3 rectangular pulses of 200 µs duration and 4 ms inter-pulse intervals were applied every 25 ms in four 20 min sessions daily. NT-3 was evaluated in supernates of homogenates from L1 – L2 and L3 – L6 segments of the spinal cord and in the soleus muscles with ELISA. In intact rats (n=4) NT-3 concentration amounted to 225 pg/mg of protein in the soleus muscle and about 60 pg/mg in lumbar segments of the spinal cord. NT-3 increased by 77% in the soleus muscle on stimulated and by 18% on non-stimulated side, comparing to intact rats. In L3 – L6 segments of the spinal cord the NT-3 was raised by 35% and 15 % on stimulated and non-stimulated side, respectively. In L1 – L2 segments there was bilateral increase of NT-3 by about 30%. We show that high-frequency low-threshold stimulation of the tibial nerve, by means of chronically implanted cuff electrodes, is capable to activate NT-3 protein both in the soleus muscle and in the caudal lumbar spinal cord indicating that also NT-3 expression is regulated in activity-dependent manner. Supported by MSE grant N N401 0480 33.
2

Does low-threshold electrical stimulation of muscle afferent fibers affect expression of the pool of neurotrophin 3 in the lumbar spinal cord of the rat?

100%
Gajewska O., Piasecka J., Kasicki S., Skup M., Czarkowska-Bauch J.
Acta Neurobiologiae Experimentalis
|
2011
|
tom 71
|
nr 1
Availability of brain-derived neurotrophic factor (BDNF) and neurotrophin 4 (NT-4) in the nervous system depends on the neuronal activity. We have previously shown that moderate locomotor exercise causes an increase of BDNF and NT-4 proteins but not neurotrophin 3 (NT-3) in the lumbar spinal cord. The questions arise whether NT-3 is regulated in a different way than BDNF and NT-4 or, that proprioceptive stimulation during treadmill locomotion is not sufficient to activate NT-3? To verify the latter possibility we applied direct electrical stimulation of the tibial nerve to activate the low-threshold muscle afferent fibers eliciting monosynaptic Hoffmann (H) reflex, an analog of the stretch reflex. Both the H-reflex and direct motor response (M), recorded from the soleus muscle, allowed keeping the strength of stimulus near the threshold of M response, confirming that stimulus is primarily addressed to lowthreshold afferents. The cuff stimulating electrode over the nerve and recording intramuscular electrodes were implanted bilaterally. The tibial nerve was stimulated unilaterally, throughout one and four weeks, starting two weeks after implantation of electrodes. The contralateral limb served as a control. Two sessions of stimulation daily, 30 min each (rectangular pulse of 300 µs duration at 0.33 Hz) were separated by about 2 h break. The total number of stimuli delivered was about 800 per day. The expression of NT-3 in the spinal cord was evaluated immunohistochemically (IR, Santa Cruz antibody) in the lumbar L4/L5 segments. Both neuropil and numerous cell bodies expressed NT-3. Perikaryonal expression was predominantly observed in the lower dorsal horn laminae (III- VI), in the intermediate zone and in the lamina IX. However, the effects of implantation and stimulation on the expression of NT-3 was negligible. The effect will be verified quantitatively with Elisa method. Supported by MSE grant N N401 0480 33.
3

Spinal cord sortilin distribution and levels change in region-specific mode after spinalization: p75 complexing in motoneurons?

86%
Gajewska O., Mankovskaya T., Ziemlinska E., Korczynski J., Czarkowska-Bauch J., Skup M.
Acta Neurobiologiae Experimentalis
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2009
|
tom 69
|
nr 3
Membrane receptor sortilin is involved in sorting and processing of proteins. In complex with p75 receptor binds proneurotrophins what can lead to apoptosis and remodeling of neuronal network. To study sortilin involvement in spinal cord (SC) remodeling after injury, we characterized its distribution patterns, levels and relation to p75 expression in L3/L4 segments, 6 weeks after SC transection at low thoracic level. In intact rats sortilin immunoreactivity (IR) was widely distributed in white and grey matter. The strongest IR was observed in glial cells of lateral and ventral funiculi. In the grey matter sortilin IR appeared in number of neurons and glial cells, except for the lamina 2, where it rarely occurred. In contrast, p75 was limited to the bundle of strongly IR fi bers in dorso-lateral lamina 2. Sortilin and p75 did not colocalise there and overlapped in isolated large neurons of lamina 9. Spinalization caused a 5% decrease of sortilin IR in the white matter not accompanied by p75 IR changes. In the grey matter sortilin IR level was not changed but frequency of p75/sortilin IR overlapping increased in lamina 9 neurons. This result indicates postlesion increase of p75/sortilin complexing in motoneurons which may refl ect activation of proneurotrophin-mediated dysfunction. Our data show lack of such interaction in primary sensory afferents. Sortilin IR in numerous cells devoid of p75 labeling confi rms that it plays also other roles. Supported by MSE P-N/029/2006 grant.
4

Locomotor exercise of spinal rats causes an increase in size of cholinergic terminals in extensor motor nuclei

72%
Czarkowska-Bauch J., Mankovskaya T., Zaremba M., Gajewska O., Ziemlinska E., Platek R., Mlodkowski M., Korczynski J., Skup M.
Acta Neurobiologiae Experimentalis
|
2009
|
tom 69
|
nr 3
Spinal cord transection causes dramatic, sustained decrease of vesicular acetylcholine transporter VAChT in terminals contacting motoneurons, as reported by Kitzman (2006, Exp Neurol 197). Cholinergic projection is known to regulate excitability of motoneurons during locomotion. The question arises if locomotor exercise of spinal, paraplegic animals might restore the role of this projection. Three groups of adult rats were tested: intact control (n=6), spinal (n=7) and spinal trained subjected to 5 weeks of treadmill locomotor training (n=8). Animals were spinalized at low thoracic segments. Gastrocnemius/soleus and anterior tibial motoneurons were prelabeled with fl uorescent dyes (FG, DY), injected to the muscles. VAChT immunoreactivity (IR) was detected using polyclonal Sigma antibody. We have found that the spinal cord transection caused a decrease of VAChT IR in boutons synapsing on cell bodies and proximal dendrites of motoneurons in L3 and L4 segments compared with that of intact rats. Surprisingly, training caused its further decay. Mean number of VAChT IR boutons did not differ consistently between groups. However, in the extensor motoneuron pools of trained animals the number of bigger VAChT IR boutons was clearly higher than in spinal non-trained and intact animals. The latter effect is in line with functional improvement of spinal trained animals, which is the most prominent in the support phase of locomotion. Supported by MSE P-N/029/2006 and N N401 0480 33 grants.
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