Paired associative stimulation improves synaptic plasticity and functional outcomes after cerebral ischemia

Paired associative stimulation is a relatively new non-invasive brain stimulation technique that combines transcranial magnetic stimulation and peripheral nerve stimulation. The effects of paired associative stimulation on the excitability of the cerebral cortex can vary according to the time interval between the transcranial magnetic stimulation and peripheral nerve stimulation. We established a model of cerebral ischemia in rats via transient middle cerebral artery occlusion. We administered paired associative stimulation with a frequency of 0.05 Hz 90 times over 4 weeks. We then evaluated spatial learning and memory using the Morris water maze. Changes in the cerebral ultra-structure and synaptic plasticity were assessed via transmission electron microscopy and a 64-channel multi-electrode array. We measured mRNA and protein expression levels of brain-derived neurotrophic factor and N-methyl-D-aspartate receptor 1 in the hippocampus using a real-time polymerase chain reaction and western blot assay. Paired associative stimulation treatment significantly improved learning and memory in rats subjected to cerebral ischemia. The ultra-structures of synapses in the CA1 area of the hippocampus in rats subjected to cerebral ischemia were restored by paired associative stimulation. Long-term potentiation at synapses in the CA3 and CA1 regions of the hippocampus was enhanced as well. The protein and mRNA expression of brain-derived neurotrophic factor and N-methyl-D-aspartate receptor 1 increased after paired associative stimulation treatment. These data indicate that paired associative stimulation can protect cog-nition after cerebral ischemia. The observed effect may be mediated by increases in the mRNA and protein expression of brain-derived neurotrophic factor and N-methyl-D-aspartate receptor 1, and by enhanced synaptic plasticity in the CA1 area of the hippocampus. The animal experiments were approved by the Animal Ethics Committee of Tongji Medical College, Huazhong University of Science & Technology, China(approval No. TJ-A20151102) on July 11, 2015.

Effects of Repetitive Transcranial Magnetic Stimulation on Astrocytes Proliferation and nNOS Expression in Neuropathic Pain Rats

This study investigated the effects of different frequencies of repetitive transcranial magnetic stimulation （rTMS） on chronic neuropathic pain in rats. The behavior of rats with experimental chronic neuropathic pain was observed, and the expression of neuronal nitric oxide synthase （nNOS） in the ipsilateral dorsal root ganglions （DRGs） and the activation and proliferation of astrocytes in the ipsilateral spinal dorsal horn were detected. Thirty-two male Sprague-Dawley rats were randomly divided into four groups： sham-operated group, sham-rTMS group, 1 Hz group and 20 Hz group （8 rats in each group）. Chronic constriction nerve injury induced by sciatic nerve ligation was made to establish the models of the chronic neuropathic pain in rats except those in the sham-operated group. Then we applied different frequencies of rTMS to the primary motor cortex （M1） contralateral to the pain side once daily for 10 consecutive days. Pain behavior scores were observed before and after treatment. Western blot analysis was used to detect the expression of nNOS in ipsilateral L4-6 DRGs. Double immunofluorescent labeling for glial fibrillary acidic protein （GFAP） and 5-bromo-2- deoxyuridine （BrdU） was employed to observe the activation and proliferation of astrocytes in the ipsilateral L4-6 spinal dorsal horn. After rTMS treatment, the spontaneous pain behavior scores were significantly lower in the 20 Hz group than those in the sham-rTMS group （P〈0.05）. Moreover, the brush-evoked pain behavior scores were significantly lower in the 20 Hz group than those in the sham-rTMS and 1 Hz group （P〈0.05）, suggesting that the spontaneous pain and brush-evoked pain in the 20 Hz group were significantly alleviated. Western blot analysis revealed that the expression of nNOS in ipsilateral L4-6 DRGs was significantly decreased in the 20 Hz group as compared with the sham-rTMS group and the 1 Hz group （P〈0.01） after rTMS treatment. Double immunofluorescence suggested that the expression of GFAP and the co-localization with BrdU in astrocytes were less in the sham-operated group than those in the sham-rTMS group and the 1 Hz group in L4-6 spinal dorsal horn ipsilateral to the neuropathic pain. After rTMS treatment, the expression of GFAP and the co-localization with BrdU decreased in the 20 Hz group as compared with the sham-rTMS group and the 1 Hz group （P〈0.05）. In addition, the alleviation degree of spontaneous pain and brush-evoked pain in the 20 Hz group was negatively correlated with the expression of nNOS in ipsilateral DRGs and the number of GFAP/BrdU co-labelled astrocytes in L4-6 spinal dorsal horn ipsilateral to the neuropathic pain （P〈0.05）. It was suggested that high-frequency rTMS may relieve neuropathic pain through down-regulating the overexpression of nNOS in ipsilateral DRGs and inhibiting the activity and proliferation of astrocytes in L4-6 spinal dorsal horn ipsilateral to the neuropathic pain.

Effect of Paired Associative Stimulation on Motor Cortex Excitability in Rats

Paired associative stimulation (PAS),combining transcranial magnetic stimulation (TMS) with electrical peripheral nerve stimulation (PNS) in pairs with an optimal interstimulus interval (ISI)in between,has been shown to influence the excitability of the motor cortex (MC)in humans.However,the underlying mechanisms remain unclear.This study was designed to explore an optimal protocol of PAS,which can modulate the excitability of MC in rats,and to investigate the underlying mechanisms. The resting motor thresholds (RMTs) of TMS-elicited motor evoked potentials (MEPs) recorded from the gastrocnemius muscle and the latency of P1 component of somatosensory evoked potentials (SEPs) induced by electrical tibial nerve stimulation were determined in male Sprague-Dawley rats (n=10).Sixty rats were then randomly divided into 3 groups:a PAS group (further divided into 10 subgroups at various ISIs calculated by using the latency of P1,n=5,respectively),a TMS (only)group (n=5)and a PNS (only)group (n=5).Ninety repetitions of PAS,TMS and PNS were administered to the rats in the 3 groups,respectively,at the frequency of 0.05 Hz and the intensity of TMS at 120% RMT and that of PNS at 6 mA.RMTs and motor evoked potentials'amplitude (MEPamp)were recorded before and immediately after the interventions.It was found that the MEPamp significantly decreased after PAS at ISI of 5 ms (P<0.05),while the MEPamp significantly increased after PAS at ISI of 15ms,as compared with those before the intervention (P<0.05).However,the RMT did not change significantly after PAS at ISI of 5 ms or 15 ms (P>0.05).PAS at other ISis as well as the sole use of TMS and PNS induced no remarkable changes in MEPamp and RMT.In conclusion,PAS can influence motor cortex excitability in rats.Neither TMS alone nor PNS alone shows significant effect.

Effects of combined rTMS and visual feedback on the rehabilitation of supernumerary phantom limbs in a patient with spinal cord injury:A case report

BACKGROUND Supernumerary phantom limb(SPL)caused by spinal cord injury(SCI)has previously been reported in several studies.However,the mechanisms and management of SPL in SCI patients are still not fully understood.Herein,we report a rare case of SPL in a patient with incomplete SCI.CASE SUMMARY A 46-year-old man complained of four hands 7 d after SCI.He was diagnosed with SPL complicated with actual limb neuropathic pain.Following a period of treatment with neurotrophic agents and Chinese traditional and analgesic medications,SPL symptoms and actual limb pain did not improve.However,his symptoms gradually lessened after combined treatment with high-frequency repetitive transcranial magnetic stimulation(rTMS),a promising neuromodulation technique,over the M1 cortex and visual feedback.After 7 wk of this treatment,SPL disappeared completely and actual limb pain was significantly relieved.CONCLUSION Cerebral plasticity changes may be a mechanism underlying the occurrence of non-painful SPL in SCI patients,and high-frequency rTMS applied to the M1 cortex could be a promising treatment method for SPL.